Patent Publication Number: US-2019170614-A1

Title: Analysis Cartridge and Method for Using Same

Description:
CROSS REFERENCE TO RELATED APPLICATIONS 
     This application claims the benefit of U.S. Provisional Application No. 62/156,441, filed May 4, 2015, U.S. Provisional Application No. 62/149,988, filed Apr. 20, 2015, and U.S. Provisional Application No. 62/018,448, filed Jun. 27, 2014, which are all incorporated by reference herein in their entireties. 
    
    
     FIELD OF THE INVENTION 
     The present invention is directed to the field of carbonyl detection and quantitation, and in particular the detection and quantitation of the concentration of carbonyl containing moieties in biological samples. 
     BACKGROUND OF THE INVENTION 
     The detection of carbonyl containing moieties is known but the precise detection of specific low concentrations of specific carbonyl containing moieties in biological samples is not known. The use of carbonyl&#39;s to induce the polymerization of o-phenylene diamine and p-phenylene diamine at high temperature is known to produce solid polymers for subsequent use in manufacturing products, but the use of phenylene diamine derivatives is not known to be used in methods to detect carbonyl containing moieties in a number of biological samples. In addition, measuring the fluorescence of a fluorogenic species in solution to determine the presence of molecules corresponding to the species is known, as well as the quantitation of the concentration of such molecules in a given sample. In addition, breath analysis devices for measuring alcohol levels are known. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  is a perspective view of the breath analysis system in accordance with a preferred embodiment of the present invention with the door opened to show the analysis cartridge in the pocket; 
         FIG. 2  is a cross-sectional elevational view of the analysis cartridge; 
         FIG. 3A  is a cross-sectional perspective view of the analysis cartridge; 
         FIG. 3B  is a cross-sectional exploded perspective view of the analysis cartridge; 
         FIG. 4  is an exploded view of the handle assembly; 
         FIG. 5A  is a cross-sectional perspective view of the analysis cartridge prior to being connected to the handle assembly; 
         FIG. 5B  is a cross-sectional perspective view of the analysis cartridge after being connected to the handle assembly; 
         FIG. 6A  is a perspective view of the analysis cartridge prior to being connected to the handle assembly: 
         FIG. 6B  is a perspective view of the analysis cartridge connected to the handle assembly; 
         FIG. 7  is a bottom plan view of the analysis cartridge; 
         FIG. 8  is a perspective view of the back of the analysis device; 
         FIG. 9  is a perspective view of the back of the analysis device with the battery cover removed; 
         FIG. 10  is a perspective view of the analysis device with half of the case removed; 
         FIG. 11  is an exploded perspective view of the analysis device; 
         FIG. 12  is a top perspective view of the rotation assembly with the analysis cartridge in the pocket; 
         FIG. 13  is an exploded perspective view of the rotation assembly; 
         FIG. 14  is another perspective view of bottom perspective view of the rotation assembly; 
         FIG. 15  is a perspective view of the rotation assembly with the analysis cartridge in the pocket; 
         FIG. 16  is a perspective view of the rotatable portion with the analysis cartridge in the pocket; 
         FIG. 17A  is an exploded perspective view of the rotatable portion with the analysis cartridge in the pocket; 
         FIG. 17B  is another exploded perspective view of the rotatable portion; 
         FIG. 17C  is an exploded perspective view of the optical system; 
         FIG. 17D  is a plan view of the bottom half of the optical system housing; 
         FIG. 17E  is a perspective view of the optical system; 
         FIG. 18  is a perspective view of the second fixed member that includes the cam track; 
         FIG. 19A  is a perspective view of the rotation assembly with the arm in the stowed position; 
         FIG. 19B  is a perspective view of the rotation assembly with the arm in the deployed position; 
         FIG. 20  is a perspective view of a portion of the rotatable assembly with the second halve of the housing removed to show the components of the optical system; 
         FIG. 21A  is a cross-sectional end view of the rotation assembly showing the rotatable portion in the first position (also referred to as the start position); 
         FIG. 21B  is a cross-sectional end view of the rotation assembly showing the rotatable portion in the second position (also referred to as the first mixing position); 
         FIG. 21C  is a cross-sectional end view of the rotation assembly showing the rotatable portion in the third position (also referred to herein as the baseline reading position); 
         FIG. 21D  is a cross-sectional end view of the rotation assembly showing the arm in a stowed position and the rotatable portion rotating toward the fourth position; 
         FIG. 21E  is a cross-sectional end view of the rotation assembly showing the rotatable portion in a fourth position (also referred to as the insertion position) and the arm in the deployed position; 
         FIG. 22  is a cross-sectional end view of the rotation assembly showing the rotatable portion in the fifth position (also referred to as the analysis position); 
         FIG. 23  is a cross-sectional end view of the rotation assembly showing the rotatable portion in the sixth position, where the analysis cartridge can be removed; 
         FIG. 24  is an exploded perspective view of an analysis cartridge system that includes a breath analysis cartridge and a fluorescence analysis cartridge in accordance with another preferred embodiment of the present invention: 
         FIG. 25  is a cross-sectional view of the breath analysis cartridge of  FIG. 24  with the ampule assembly in elevation; 
         FIG. 25A  is a cross-sectional view of the ampule assembly of the breath analysis cartridge; 
         FIG. 26  is a cross-sectional view of the breath analysis cartridge of  FIG. 24  with the ampule assembly in elevation and the ampule member pushed in; 
         FIG. 26A  is a cross-sectional view of the ampule assembly of the breath analysis cartridge; 
         FIG. 27  is a cross-sectional view of the fluorescence analysis cartridge of  FIG. 24 ; 
         FIG. 28  is an elevational view of the analysis cartridge system of  FIG. 24  with the breath analysis cartridge received on the fluorescence analysis cartridge; 
         FIG. 29  is a cross-sectional view of the analysis cartridge system of  FIG. 24 ; 
         FIG. 30  is a cross-sectional view of an analysis cartridge in accordance with another preferred embodiment of the present invention; 
         FIG. 31  shows graphs depicting the emission spectrum of the reaction of mPDA with 1-hexanal as a function of time; 
         FIG. 32  shows a graph depicting the increase in fluorescence over time of the reaction of mPDA with 1-hexanal being the carbonyl containing moiety; 
         FIG. 33A  shows a graph depicting the increase in fluorescence over time of the reaction with 1-hexanal as a function of sodium dodecyl sulfate (“SDS”) concentration from 0.01 to 0.4% (w/v); 
         FIG. 33B  shows a graph depicting the increase in fluorescence over time of reaction with 1-hexanal as compared to a blank, with SDS concentration at 0.2% SDS; 
         FIG. 33C  shows a graph depicting the increase in fluorescence over time of the reaction with 1-hexanal as compared to a blank, with SDS concentration at 0.4% SDS; 
         FIG. 34  shows a graph displaying fluorescence as a function of 1-hexanal concentration; 
         FIG. 35  shows a chart depicting the relative fluorescence as a function of aldehyde chain length; and 
         FIG. 36  shows a chart depicting the relative fluorescence of selected small aromatic amines. 
     
    
    
     SUMMARY OF THE PREFERRED EMBODIMENTS 
     In accordance with an aspect of the present invention there is provided an analysis cartridge that includes a main body portion having an upper portion that defines an upper chamber and a lower portion that defines a fluid chamber, and a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough. In the first position, the opening partially defines the upper chamber and in the second position the opening partially defines the fluid chamber. In a preferred embodiment, the filter assembly is movable within a cylindrical sleeve that extends from the upper chamber to the fluid chamber. Preferably, the sleeve includes a top opening such that an upper surface of the filter assembly is exposed to an exterior of the main body portion. 
     In a preferred embodiment, the filter assembly includes a cylindrically shaped filter holder that includes the opening extending transversely therethrough and two filters positioned such that they span the opening. The filters define a substrate space therebetween and a substrate is disposed in the substrate space. In a preferred embodiment, the substrate is silica and the fluid chamber includes an elution solution or rinse therein. Preferably, the upper chamber is a breath chamber that includes a breath entry opening, a breath exit opening and a breath path therebetween. In a preferred embodiment, the analysis cartridge includes a pressure measurement hole defined in a wall of the upper portion that communicates the breath chamber with a pressure tunnel that extends through the main body portion and to a pressure recess defined in the lower portion. 
     In a preferred embodiment, a phenylene diamine derivative is disposed in the analysis cartridge. Preferably, the analysis cartridge includes an ampule member having a fluorescence chromophore space with the phenylene diamine derivative disposed therein and the fluid chamber includes an elution solution disposed therein. The ampule member is movable between a first position where the phenylene diamine derivative is separated from the elution solution and a second position where the phenylene diamine derivative is disposed in the elution solution. In a preferred embodiment, the phenylene diamine derivative is m-phenylene diamine. 
     In accordance with an embodiment of the present invention there is provided a method that includes (a) obtaining an analysis cartridge that includes a main body portion having an upper portion that defines a breath chamber and a lower portion that defines a fluid chamber, and a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough and in the first position the opening partially defines the upper chamber and in the second position the opening partially defines the fluid chamber and the fluid chamber includes an elution solution therein, as described below. The method also includes (b) capturing a breath sample in the filter assembly, (c) moving the filter assembly from the first position to the second position, and (d) eluting constituents of the breath sample into the elution solution to form a constituent solution. In a preferred embodiment, prior to step (c) the method includes inserting the analysis cartridge into an analysis pocket, and rotating the analysis cartridge to an insertion position where an arm performs step (c). 
     In accordance with another embodiment of the present invention there is provided an analysis cartridge system that includes a breath analysis cartridge and a fluorescence analysis cartridge. The breath analysis cartridge includes a main body portion that includes an upper portion that defines a breath chamber and a lower portion that defines a fluid chamber. The breath chamber includes a breath entry opening, a breath exit opening and a breath path therebetween, and the lower portion includes a receiver member extending therefrom. The breath analysis cartridge also includes a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough, and, in the first position, the opening partially defines the breath chamber and is part of the breath path and in the second position the opening partially defines the fluid chamber. The fluorescence analysis cartridge includes a main body portion that includes an upper portion that defines an upper chamber and a lower portion that defines a fluid chamber. The upper chamber includes a front opening that is adapted to receive the receiver member of the breath analysis cartridge therein. The fluorescence analysis cartridge also includes a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough, and, in the first position, the opening partially defines the upper chamber and in the second position the opening partially defines the fluid chamber. In a preferred embodiment, the in both the breath analysis cartridge and the fluorescence analysis cartridge the breath or upper chamber is sealed from the fluid chamber when the filter assembly is in the first position. 
     In a preferred embodiment, the breath analysis cartridge includes an ampule member that is slideable within a slide tube between the first and second positions. Preferably, the ampule member includes at least one opening therein that is sealed from fluid communication with the fluid chamber when the ampule member is in the first position, and is in fluid communication with the fluid chamber when the ampule member is in the second position. In a preferred embodiment, the filter assembly divides the upper chamber of the fluorescence analysis cartridge into a front chamber and a rear chamber. The front chamber includes a piercing member disposed therein that is adapted to pierce a breakable barrier of the ampule member. Preferably, the rear chamber of the fluorescence analysis cartridge includes an absorption member positioned therein. 
     In a preferred embodiment, the breath analysis cartridge includes a removable mouthpiece that defines a central opening that is in communication with the breath chamber. The mouthpiece includes a sleeve portion that is received in the breath entry opening and a mouthpiece portion. Preferably, the mouthpiece includes a stopper that abuts the main body portion. The stopper includes an alignment member extending therefrom that is received in an alignment opening in the main body portion. In a preferred embodiment, the fluorescence analysis cartridge includes opposing light entry and light exit windows positioned on opposite sides of the fluid chamber, and a fluorescence window positioned on the bottom of the main body portion. Preferably, the light entry and light exit windows each include an outer surface, and wherein the outer surfaces are parallel to one another. Preferably, the fluorescence window includes an outer surface and the outer surface of the fluorescence window is perpendicular to the outer surface of the light entry window. 
     In accordance with another embodiment of the present invention there is provided a method that includes obtaining an analysis cartridge system that includes a biological analysis cartridge and an identified constituent analysis cartridge. The biological analysis cartridge has an upper chamber and a fluid chamber, and the identified constituent analysis cartridge has an upper chamber and a fluid chamber. The method also includes capturing a biological sample as described below on a substrate as described below positioned in the upper chamber in the biological analysis cartridge, moving the substrate from the upper chamber to the fluid chamber, which includes a first elution solution therein as described below, eluting constituents of the biological sample into the first elution solution to form a constituent solution as described below, releasing a moiety into the second solution to form a first identifiable constituents solution, transferring the first identifiable constituents solution as described below to the upper chamber of the identified constituent analysis cartridge, such that identified constituents are captured on a substrate positioned in the upper chamber, moving the substrate from the upper chamber to a fluid chamber that includes a second elution solution therein as described below, and eluting the identified constituents into the second elution solution to form a second identifiable constituents solution as described below. 
     In a preferred embodiment, the biological analysis cartridge is a breath analysis cartridge, the identified constituent analysis cartridge is a fluorescence analysis cartridge, and the biological sample is a breath sample. Preferably, the moiety is a fluorescence chromophore as described below. 
     In accordance with another embodiment of the present invention there is provided a method of forming a solution within a breath analysis cartridge that includes a main body portion with an upper portion that defines a breath chamber, a lower portion that defines a fluid chamber having an elution solution disposed therein and an ampule member that is movable between a first position and a second position. The ampule member includes a fluorescence chromophore space having a fluorescence chromophore disposed therein. The method includes moving the ampule member from the first position where the fluorescence chromophore space and fluorescence chromophore are separated from the fluid chamber to the second position where the fluorescence chromophore space is in communication with the fluid chamber, and mixing the fluorescence chromophore with the elution solution. 
     In accordance with another embodiment of the present invention there is provided an analysis cartridge that includes a main body portion that includes an upper portion that defines a breath chamber and a lower portion that defines a fluid chamber. The breath chamber includes a breath entry opening, a breath exit opening and a breath path therebetween. The analysis cartridge also includes a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough, and, in the first position, the opening partially defines the breath chamber and is part of the breath path and in the second position the opening partially defines the fluid chamber. In a preferred embodiment, the filter assembly includes first and second filters positioned in the opening and the first and second filters define a substrate space therebetween with a substrate disposed therein. Preferably, the substrate is incorporated with an active reactive capture agent. In a preferred embodiment, the active reactive capture agent is a fluorescent hydrazine or aminooxy compound. 
     In accordance with another embodiment of the present invention there is provided a method of forming a fluorescing solution within an analysis cartridge that includes a main body portion with an upper portion that defines a breath chamber, a lower portion that defines a fluid chamber having an elution solution disposed therein and a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough, and, in the first position, the opening partially defines the breath chamber and in the second position the opening partially defines the fluid chamber. The filter assembly includes a substrate incorporated with an active reactive capture agent disposed therein. The method includes capturing carbonyl containing moieties on the substrate, moving the filter assembly from the first position to the second position, and eluting the carbonyl containing fluorescence chromophores and active reactive capture agent into the elution solution to form the fluorescing solution. 
     In accordance with another embodiment of the present invention there is provided a breath capture assembly that includes a handle assembly having an elongated main body portion that defines a handle interior, a cap disposed at an end of the main body portion that includes a pressure opening defined therein, and a pressure transducer disposed in the handle interior. The breath capture assembly also includes an analysis cartridge received on an upper end of the handle assembly. The analysis cartridge includes a main body portion that has an upper portion that defines a breath chamber and a lower portion that defines a fluid chamber. The breath chamber includes a breath entry opening, a breath exit opening and a breath path therebetween. The analysis cartridge includes a filter assembly that is movable along a filter assembly path between a first position and a second position. The filter assembly has an opening defined therethrough, and, in the first position, the opening partially defines the breath chamber and is part of the breath path and in the second position the opening partially defines the fluid chamber. 
     In a preferred embodiment, the pressure measurement hole is defined in a wall of the upper portion of the analysis cartridge and communicates the breath chamber with a pressure tunnel that extends through the main body portion. A pressure path is defined from the breath chamber, through the pressure measurement hole, the pressure tunnel, the pressure opening and to the pressure transducer. Preferably, the cap of the handle assembly includes a pressure protrusion extending upwardly therefrom that is sealingly received in a pressure recess in the analysis cartridge. The pressure recess is in communication with the pressure tunnel, and the pressure opening is defined in the pressure protrusion. In a preferred embodiment, the cap includes a seat defined therearound, and a collar depending downwardly from the analysis cartridge is received on the seat. The cap preferably includes an attachment protrusion extending radially outwardly therefrom that is received in an attachment recess defined in the collar of the analysis cartridge. 
     In a preferred embodiment, a hollow extension extends downwardly from the cap of the handle assembly and into the handle interior. The hollow extension is part of the pressure path. Preferably, a pressure tube is received on the hollow extension and is in the pressure path between the hollow extension and the pressure transducer. 
     In a preferred embodiment, the analysis cartridge includes a breakable barrier disposed between the breath chamber and the fluid chamber when the filter assembly is in the first position to seal the breath chamber from the fluid chamber. 
     In accordance with another embodiment of the present invention there is provided an analysis device that includes a case defining a case interior, a door movable between an open and a closed position, and a rotation assembly positioned in the case interior that includes first and second fixed members and a rotatable portion positioned between the first and second fixed members. The rotatable portion is rotatable about a rotation axis with respect to the first and second fixed members. The rotatable portion includes a shroud that has a funnel portion defined therein for receiving an object to be rotated. The shroud includes a pocket opening defined at the top thereof and the rotation assembly includes a fluorescence detection assembly positioned generally below the shroud. The analysis device also includes a motor that drives rotation of the rotatable portion and a controller that controls the motor and the fluorescence detection assembly. 
     In a preferred embodiment, the shroud includes a pocket opening and an analysis opening opposite to one another, and the shroud includes walls that taper between the pocket opening and the analysis opening. The fluorescence detection assembly preferably includes a housing that has an analysis cartridge receiving portion with a well defined therein that is aligned with the analysis opening in the shroud to form an analysis pocket. In a preferred embodiment, the analysis cartridge receiving portion cooperates with the shroud to define a light entry aperture, a light exit aperture and a fluorescence aperture. Preferably, the fluorescence detection assembly includes a light that is configured to be directed along a light path that extends through a light chamber defined in the housing, through the light entry aperture, through the light exit aperture, and into a light trap. 
     In a preferred embodiment, the fluorescence detection assembly includes a detector for receiving fluorescence emitted through the fluorescence opening and through a fluorescence chamber defined in the housing. Preferably, the fluorescence chamber is generally orthogonal to the light chamber. In a preferred embodiment, the analysis device includes an arm that is pivotal between a stowed position and a deployed position. The arm includes a first end that extends through an arm opening defined in the shroud when in the deployed position. When the rotatable portion rotates from a start position to an insertion position the arm pivots from the stowed position to the deployed position. In a preferred embodiment, the arm is biased toward the stowed position and includes a second end that is operationally associated with a cam surface on the second fixed member. The cam surface preferably has a stowed end that is associated with the stowed position of the arm and a deployed end that is associated with the deployed position of the arm and includes an increasing radius from the stowed end to the deployed end. In a preferred embodiment, the arm includes a ball bearing on the second end thereof that interacts with the cam surface. Preferably, the arm is pivotal on a shaft that extends from the shroud. 
     In accordance with another embodiment of the present invention there is provided a rotation assembly that includes first and second fixed members, and a rotatable portion positioned between the first and second fixed members that is rotatable about a rotation axis with respect to the first and second fixed members. The rotatable portion includes a shroud that has a funnel portion defined therein for receiving an object to be rotated and an arm that is pivotal between a stowed position and a deployed position. The arm includes a first end that extends through an arm opening defined in the shroud when in the deployed position. The analysis device also includes a motor that drives rotation of the rotatable portion. When the rotatable portion is rotated from a start position to an insertion position the arm pivots from the stowed position to the deployed position. In a preferred embodiment, the shroud includes a pocket opening and an analysis opening opposite to one another and walls that taper between the pocket opening and the analysis opening. 
     In a preferred embodiment, the shroud includes first and second axle members extending outwardly therefrom that are received in opening in the first and second fixed members, respectively. Preferably, the shroud includes at least one internally threaded fastener receiver member extending therefrom. The housing of the fluorescence detection assembly includes at least one receiver tube and a threaded receiver extends through the receiver tube and into the fastener receiver member to secure the shroud to the housing. Preferably, the housing includes first and second housing halves. A first receiver tube is located on the first housing half and a second receiver tube is located on the second housing half. The threaded receiver extends through the first and second receiver tubes and into the fastener receiver member to secure the shroud to the housing. 
     In accordance with another embodiment of the present invention there is provided a handle assembly for use with a breath analysis system that includes an analysis cartridge and an analysis device. The handle includes an elongated main body portion that defines a handle interior, a cap disposed at an end of the main body portion that includes a pressure opening defined therein, a pressure transducer disposed in the handle interior, and a pressure path defined between the pressure opening and the pressure transducer. In a preferred embodiment, the cap includes a pressure protrusion extending upwardly therefrom and the pressure opening is defined in the pressure protrusion. Preferably, the handle interior includes a magnet disposed therein that interacts with a magnet in the analysis device. The magnet is positioned in a magnet recess defined in the cap. 
     In accordance with another embodiment of the present invention there is provided a filter assembly that includes a main body portion having a generally cylindrical shape that defines a first axis, an opening defined transversely through the main body portion that extends generally perpendicularly to the first axis, first and second filters spanning the opening and defining a substrate space therebetween, and a substrate disposed in the substrate space. Preferably, the first and second filters comprise a plastic having pores defined therethrough. In a preferred embodiment, the main body portion includes guide rails on an outside surface thereof that extend generally parallel to the axis. Preferably, the main body portion includes a lower surface that includes at least one piercer extending downwardly therefrom. 
     In accordance with another embodiment of the present invention there is provided a method of making a filter assembly that includes obtaining a filter holder having a main body portion with a generally cylindrical shape that defines a first axis and includes an opening defined transversely through the main body portion that extends generally perpendicularly to the first axis, dosing a first filter with a substrate, pressing a second filter onto the substrate, and positioning the first filter, substrate and second filter into the opening such that the first and second filter span the opening. The first and second filters and substrate can be positioned in the opening together or separately. 
     In accordance with another embodiment of the present invention there is provided a fluorescence detection assembly that includes an emitter, a detector, a housing that defines an light chamber, a fluorescence chamber and a well, a light path that extends from the emitter, through the light chamber and through the well, and a fluorescence path that extends from the well, through the fluorescence chamber and to the detector. In a preferred embodiment, the fluorescence detection assembly includes a first lens and a first filter positioned within the light path. Preferably, the fluorescence detection assembly includes second lens and a second filter positioned within the fluorescence path. In a preferred embodiment, the fluorescence detection assembly includes at least one of a first light baffle positioned within the light path between the emitter and the first lens, a second light baffle positioned within the light path between the first lens and the first filter and a third light baffle positioned within the light path between the first filter and the well. The first baffle includes a first light baffle aperture defined therein that has a smaller inner diameter than an inner diameter of the light chamber. The second baffle includes a second light baffle aperture defined therein that has a smaller inner diameter than the inner diameter of the first light baffle aperture. The third baffle includes a third light baffle aperture defined therein that has a smaller inner diameter than the inner diameter of the second light baffle aperture. 
     In a preferred embodiment, the light trap is positioned at a distal end of the light path and includes a first wall that is angled between about 25° and about 45° with respect to the light path. Preferably, the light trap includes a second wall connected to the first wall and the second wall is not perpendicular to the light path. In a preferred embodiment, the housing is comprised of an upper housing half and a lower housing half and the lower housing half includes an analysis cartridge receiving portion that defines the well. In a preferred embodiment, the upper housing half includes a flange that extends downwardly therefrom and overlaps a flange extending upwardly from the lower housing half. In a preferred embodiment, an analysis cartridge is positioned in the well that includes a light entry window, a light exit window and a fluorescence window. The light entry window and light exit window are positioned along the light path. 
     In a preferred embodiment, the housing is comprised of an upper housing half and a lower housing half that cooperate to define a first lens pocket that houses the first lens, a first filter pocket that houses the first filter, a second lens pocket that houses the second lens, and a second filter pocket that houses the second filter. Preferably, the fluorescence detection assembly includes a shroud connected to the housing that includes a pocket opening and an analysis opening opposite to one another and a funnel portion therebetween. The funnel portion cooperates with the well to define an analysis pocket and the shroud at least partially defines the light entry aperture and the fluorescence aperture. 
     In accordance with another embodiment of the present invention there is provided a method of detecting fluorescence that includes emitting light from an emitter into an light chamber and along a light path that includes a sensing chamber therealong. The sensing chamber includes a fluorescing solution therein. The emitted light passes through the fluorescence solution and produces a fluorescence light, and wherein the fluorescence light is emitted from the sensing chamber into a fluorescence chamber along a fluorescence path, and detecting a fluorescence signal of the fluorescence light. 
     In accordance with another embodiment of the present invention there is provided a method of detecting fluorescence that includes inserting an analysis cartridge into an analysis pocket. The analysis cartridge includes a filter assembly that includes a substrate having a carbonyl containing moiety thereon. The method also includes rotating the analysis cartridge from a start position to an insertion position, moving the filter assembly within the analysis cartridge from an upper chamber to a fluid chamber that contains an elution solution, rotating the analysis cartridge from the insertion position to an analysis position such that the elution solution drains through the filter assembly and the carbonyl containing moiety is eluted into the elution solution to form a fluorescing solution, and analyzing the fluorescence of the fluorescing solution. 
     In accordance with another embodiment of the present invention there is provided a breath analysis system that includes a breath capture assembly that includes a handle assembly that includes an elongated main body portion that defines a handle interior, a pressure opening defined in an end of the elongated main body portion, and a pressure transducer disposed in the handle interior. The breath analysis system also includes an analysis cartridge received on an upper end of the handle assembly. The analysis cartridge includes a main body portion that includes an upper portion that defines a breath chamber, and a lower portion that defines a fluid chamber. The breath chamber includes a breath entry opening, a breath exit opening and a breath path therebetween. The analysis cartridge includes a filter assembly that is movable along a filter assembly path between a breath capture position and an analysis position. The filter assembly has an opening defined therethrough, and, in the breath capture position, the opening partially defines the breath chamber and is part of the breath path and in the analysis position the opening partially defines the fluid chamber. The system also includes an analysis device that includes a case defining a case interior, a door movable between an open and closed position, and a rotation assembly positioned in the case interior that includes a shroud that has a funnel portion defined therein for receiving the analysis cartridge. The system also includes a controller that controls the motor and the fluorescence detection assembly. The pressure transducer is in communication with the controller. 
     In accordance with another embodiment of the present invention there is provided a method for detecting and quantifying carbonyl containing moieties in breath. The method includes (a) providing an analysis cartridge, (b) connecting the analysis cartridge to a handle assembly, (c) collecting a breath sample of carbonyl containing moieties on a filter assembly, (d) labeling the carbonyl containing moieties to provide a labeled solution, (e) inserting the labeled solution into an analysis device, (f) directing light within a predetermined wavelength range through the labeled solution, thereby producing a fluorescence, and (g) detecting the fluorescence. 
     It will be appreciated that any biological sample can be analyzed using the system. Breath constituents other than carbonyl containing moieties (CCM) or aldehydes can be captured and analyzed as desired. U.S. Patent Publication Nos. 2003/0208133 and 2011/0003395 are incorporated by reference herein in their entireties. 
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS 
     The following description and drawings are illustrative and are not to be construed as limiting. Numerous specific details are described to provide a thorough understanding of the disclosure. However, in certain instances, well-known or conventional details are not described in order to avoid obscuring the description. References to one or another embodiment in the present disclosure can be, but not necessarily are, references to the same embodiment; and, such references mean at least one of the embodiments. 
     Reference in this specification to “one embodiment” or “an embodiment” means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the disclosure. Appearances of the phrase “in one embodiment” in various places in the specification do not necessarily refer to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments. Moreover, various features are described which may be exhibited by some embodiments and not by others. Similarly, various requirements are described which may be requirements for some embodiments but not other embodiments. 
     The terms used in this specification generally have their ordinary meanings in the art, within the context of the disclosure, and in the specific context where each term is used. Certain terms that are used to describe the disclosure are discussed below, or elsewhere in the specification, to provide additional guidance to the practitioner regarding the description of the disclosure. For convenience, certain terms may be highlighted, for example using italics and/or quotation marks: The use of highlighting has no influence on the scope and meaning of a term; the scope and meaning of a term is the same, in the same context, whether or not it is highlighted. It will be appreciated that the same thing can be said in more than one way. 
     Consequently, alternative language and synonyms may be used for any one or more of the terms discussed herein. Nor is any special significance to be placed upon whether or not a term is elaborated or discussed herein. Synonyms for certain terms are provided. A recital of one or more synonyms does not exclude the use of other synonyms. The use of examples anywhere in this specification including examples of any terms discussed herein is illustrative only, and is not intended to further limit the scope and meaning of the disclosure or of any exemplified term. Likewise, the disclosure is not limited to various embodiments given in this specification. 
     Without intent to further limit the scope of the disclosure, examples of instruments, apparatus, methods and their related results according to the embodiments of the present disclosure are given below. Note that titles or subtitles may be used in the examples for convenience of a reader, which in no way should limit the scope of the disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains. In the case of conflict, the present document, including definitions, will control. 
     It will be appreciated that terms such as “front,” “back,” “top,” “bottom,” “side,” “short,” “long,” “up,” “down,” and “below” used herein are merely for ease of description and refer to the orientation of the components as shown in the figures. It should be understood that any orientation of the components described herein is within the scope of the present invention. 
       FIGS. 1-30  show a breath analysis system  10  for analyzing carbonyl containing moieties (“CCM”) in a patient&#39;s breath. As shown in  FIG. 1 , the system  10  generally includes a handle assembly  12 , an analysis cartridge  14  and an analysis device  16 . Generally, the handle assembly  12  and analysis cartridge  14  are used by the clinician and patient to capture certain components of the patient&#39;s breath (as described more fully below), and the analysis device  16  is used to analyze the captured components. 
     The analysis cartridge  14  shown in  FIGS. 2-7 and 21A-23  will now be described. In a preferred embodiment, the analysis cartridge  14  includes a main body portion  11  that includes an upper portion  29  that defines an upper or breath chamber  30  and a lower portion  31  that defines a lower or fluid chamber  32 . The breath chamber  30  includes a front opening or breath entry opening  33 , a breath exit opening  40  and a breath path P 1  therebetween. In a preferred embodiment, the breath chamber  30  tapers toward the breath exit opening  40 , however, this is not a limitation. The analysis cartridge  14  also includes a filter assembly  19  that is movable along a filter assembly path P 2  between a first or breath capture position ( FIG. 2 ) and a second or analysis position ( FIG. 21E ). The filter assembly  19  has an opening  17  defined therethrough that includes at least one and preferably two filters  26  positioned therein. In the breath capture position the opening  17  partially defines the breath chamber  30  and is part of the breath path P 1  and in the analysis position the opening  17  partially defines the fluid chamber  32 . 
     In a preferred embodiment, the analysis cartridge  14  includes a removable mouthpiece  18 , the filter assembly  19  on the top and an ampule member  22  on the bottom. As shown in  FIG. 3B , the mouthpiece  18  includes a sleeve portion  15  that is received in breath entry opening  33  on the main body portion  11 , a mouthpiece portion  18   a , a stopper  21  that abuts the main body portion  11  and an alignment member  21   a  that is received in a complementary alignment opening in the main body portion  11  (not shown). The mouthpiece  18  partially defines the breath chamber  30  and the breath path P 1 . The filter assembly  19  preferably includes two filters or frit plates  26  (sometimes referred to together as a flit stack) that are held by a frit holder  20 . The frit plates  26  span opening  17 . 
     In a preferred embodiment, the frit holder  20  includes at least one piercer  20   a  on the bottom surface thereof for piercing a breakable barrier discussed below. Preferably, the frit holder  20  includes at least one guide rail  39  on an outside surface for helping guide the frit assembly  19  as it is moved along the filter assembly path. The piercer  20   a  can be on the bottom of the guide rail  39 . Prior to use, the frit stack  26  is positioned in the breath chamber  30 . As shown in  FIG. 2 , a substrate space  27  is defined between the frit plates  26 . In a preferred embodiment, a substrate  28 , such as silica, is disposed in the substrate space  27  between the frit plates  26 . It will be appreciated that the frit plates  26  are sufficiently porous so that the breath can pass therethrough, but not so porous that the substrate  28  trapped therebetween can escape. The filters or frit plates  26  are preferably made of polyethylene spheres that are pressed and packed together in a form. When pressed together in the disc or plate shape, the spherical or roundish shape creates the voids or pores necessary for breath to get through. The spheres can be made out of different plastic materials (e.g., polyethylene, polypropylene, etc.) or teflon in different diameters. In another embodiment, the filters  26  can comprise spheres all made of the same plastic materials and of the same or different diameters. In an exemplary embodiment, the frits  26  are polyethylene or teflon frits with 10 um or 20 um pore sizes. As is described more fully below, in use, as a patient blows through the breath chamber  30 , CCM including aldehydes, collect on the substrate  28  (also referred to as CCM capture material). In a preferred embodiment, silica is used as the substrate or CCM capture material. However, this is not a limitation on the present invention and other substrates with the ability to capture CCM or aldehydes can be used. 
     In a preferred embodiment, the flit plates or filters  26  that span opening  17  are preferably press fit therein. The method of creating the filter assembly includes pressing the spherical plastic pieces into first and second filters  26 , pressing the first filter  26  into the opening  17  in the flit holder  20 . Then, the substrate  28  (preferably silica) is dosed on the first flit  26 , a second filter  26  is then pressed into the opening  17  onto the silica  28  using a predefined pressure. In another embodiment, the silica  28  can be dosed onto the first flit  26  and then the second flit  26  can be pressed onto the silica  28  to create a frit stack, prior to pressing the flit stack into the opening  17  in the flit holder  20 . In another embodiment, the filters can be disposed in grooves defined in the inside wall of the flit holder  20 . 
     As shown in  FIGS. 2-3B , the ampule member  22  comprises a main body portion  23  having a fluorescence chromophore space or trough  25  defined therein that includes an upper rim  23   a  and a lower surface  23   b . The trough  25  includes a phenylene diamine derivative (“PD derivative”)  24  disposed therein. The ampule member  22  is movable between a first position where the trough  25  and PD derivative  24  are separated from the fluid chamber  32  by a first breakable barrier  36   a  and a second position where the trough  25  is in communication with the fluid chamber  32  (where the PD derivative  24  and elution solution  34  are mixed in the fluid chamber  32 , as described below). The breakable barrier  36   a  can be a foil or the like. In a preferred embodiment, the ampule member  22  is movable within an ampule tunnel  130  that is defined in the lower portion  31  of the analysis cartridge  14 . In a preferred embodiment, the ampule member  22  includes a flange or stopper  23   c  that abuts a stopper surface  132  on the analysis cartridge  14  when the ampule member  22  is moved to the second position. The stopper  23   c  prevents the ampule member  22  from moving too far into the ampule tunnel  130  and/or into the fluid chamber  32 . In a preferred embodiment the ampule tunnel  130  is orthogonal to the fluid chamber  32 . However, this is not a limitation. 
     As shown in  FIGS. 2-3B , the fluid chamber  32  is located between the breath chamber  30  and the ampule member  22  and ampule tunnel  130 . An elution solution  34  is disposed in the fluid chamber  32 . In a preferred embodiment, the elution solution  34  includes water and ethanol, however, this is not a limitation on the present invention. In a preferred embodiment, the fluid chamber  32  is sealed from the ampule tunnel  130 . This can be done by any sealing method. In a preferred embodiment, the fluid chamber  32  is sealed from the ampule tunnel  130  by first breakable barrier  36   a . In a preferred embodiment, the fluid chamber  32  is sealed from the breath chamber  30 . This can be done by any sealing method. In a preferred embodiment, a second breakable barrier  36   b  is positioned across the filter assembly pathway P 2  (dividing the filter assembly sleeve  53 ) between the breath chamber  30  and the fluid chamber  32 . The opening between the fluid and breath chambers is referred to herein as the filter assembly opening  134  and it includes a ledge on which the second breakable barrier  36   b  is secured. The fluid chamber  32  also includes vent holes  37  to keep the elution solution  34  from getting “air locked” during mixing. 
     It will be appreciated by those of skill in the art that before use of the analysis cartridge  14  (i.e., before it is attached or connected to the handle assembly  12 ), the filter assembly  19  is in the breath capture position and the ampule member  22  is in the first position. In this configuration, the elution solution  34  in the fluid chamber  32  is separated from the filter assembly  19  in the breath chamber  30  by second breakable barrier  36   b  and the ampule member  22  in the ampule tunnel  130  by first breakable barrier  36   a.    
       FIG. 4  is an exploded view of the handle assembly  12  and the components thereof. In a preferred embodiment, the handle assembly  12  includes an elongated main body portion  101  with first and second halves  102  that define a handle interior  99  (see  FIG. 5A ), top and bottom end caps  103  and  105 , a grip  104 , a cable  106  that connects to the analysis device  16  (electric and/or data) via plug  107  and a pressure transducer  50  and associated components (circuit board  108 , pressure tube  110 , etc.). The handle assembly  12  also preferably includes a magnet  111  that interacts with a magnet  150   a  in handle storage pocket  66  and tube  150  described below. A pressure protrusion  49  extends outwardly from the upper surface  52  of the top end cap  103  (see  FIG. 5A ). A pressure opening  113  is defined in the top of the pressure protrusion  49 . In a preferred embodiment, the pressure protrusion includes an O-ring  115  therearound that seals the pressure protrusion  49  when it is coupled to the analysis cartridge  14 . Preferably, the pressure protrusion  49  is received in a pressure recess  139  (see  FIG. 7 ) defined in the lower portion  31  of the analysis cartridge. As shown in  FIGS. 5 and 5A , in a preferred embodiment, the pressure path within the handle assembly  12  extends from the pressure opening  113 , through an extension  117  on the bottom surface of the top end cap  103  (which is received in the pressure tube  110 ), through the pressure tube  110  and to the pressure transducer  50  (an end of which is received in the pressure tube  110 ). Generally, the pressure path is defined between the pressure opening  113  and the pressure transducer  50 . The cable  106  is connected to the circuit board  108 . Therefore, the pressure reading of the pressure transducer  50  can be communicated to main circuit board  74  of the analysis device  16 . 
       FIGS. 5A-6B  show the analysis cartridge  14  being attached to the handle assembly  12 . In a preferred embodiment, the analysis cartridge  14  includes a collar  138  extending downwardly from the main body portion  11 . The collar  138  includes at least one and preferably a plurality of attachment recesses  140  defined therein. One of the recesses  140  mates with an alignment or attachment protrusion  142  on the top end cap  103  of the handle assembly  12  (in another embodiment there can be more attachment protrusions  142 ). The collar  138  is also received on a seat  144  that is a part of the top end cap  103 . An annular protrusion extends outwardly from cap  103  that mates with an undercut slot on the collar  138  and that creates a snap fit. A friction fit is also within the scope of the present invention. The attachment recesses  140  allow the collar to expand when secured on the top of the handle assembly  12 . 
     The pressure recess  139  for receiving the pressure protrusion  49  on the top of the handle assembly  12  is defined within the collar  138 . The complementary attachment recess  140  and attachment protrusion  142  align the analysis cartridge  14  and handle assembly  12  during the attachment process so that the pressure protrusion  49  is received in the pressure recess  139 . The seat  144  can include a rubber material or the like for providing a friction fit with the collar  138 . Any method of attaching the analysis cartridge  14  to the handle assembly  12  is within the scope of the present invention. For example, analysis cartridge  14  to the handle assembly  12  can be connected by a threaded fit, snap fit, friction fit or the like. 
     As shown in  FIG. 5A , in the first position, the ampule member  22  extends downwardly from the lower portion  31  of the analysis cartridge  14 . Therefore, when the analysis cartridge  14  is connected to the handle assembly  12 , the lower surface  23   b  of the ampule member  22  contacts the upper surface  52  of the handle assembly  12 , thereby pushing the ampule member  22  upwardly, thus breaking the first breakable barrier  36   a , and moving the ampule member from the first position to the second position. This facilitates transferring the PD derivative  24  into the fluid chamber  32  and the elution solution  34 . It will be appreciated that the PD derivative  24  is kept separated from the elution solution  34  until the analysis cartridge  14  is connected to the handle assembly  12 . The elution solution  34  and PD derivative  24  mix to form a phenylene diamine solution (“PD solution”)  35  (further mixing of the elution solution  34  and PD derivative  24  in analysis device  16  is described below). 
     With references to  FIGS. 2-7 , another feature included in the analysis cartridge  14  is the breath pressure measurement capability. This enables a patient blowing into the analysis cartridge to know via the screen  60  on the analysis device  16  whether the blown pressure is within a predetermined range. Generally, a pressure path is defined between a pressure measurement hole  42  in the upper portion  29  of the analysis cartridge  14  (that is in communication with the breath chamber  30 ) and the pressure transducer  50  in the handle assembly  12 . As shown in  FIG. 3A , the pressure path extends from the pressure measurement hole  42  to a pressure channel  44  that extends partially around the filter assembly  19 , and to a pressure tunnel  46  that extends downwardly through the main body portion  11 . It will be appreciated that  FIG. 3A  shows a top ring cover  47  omitted from the analysis cartridge  14 . The top ring cover  47  (or other wall or barrier) encloses the pressure channel  44  to maintain the pressure. 
       FIG. 7  shows the bottom of the analysis cartridge  14  and the end of the pressure tunnel  46  that communicates with the pressure recess  139 . When the pressure protrusion  49  is received in the pressure recess  139 , the pressure tunnel  46  is communicated with the pressure opening  113 . Therefore, the complete pressure path extends from the pressure measurement hole  42 , through the pressure channel, through the pressure tunnel, through the pressure opening, through the hollow extension  117 , through the pressure tube  110  and to the pressure transducer  50 . 
     When a patient blows through the breath chamber  30  (and the frit plates  26 ), a pressure measurement is taken. In a preferred embodiment, this requires a pressure differential flow calculation. As breath is being blown through the breath chamber, depending on how hard the person is blowing, there is a pressure differential that is created in the distal space  38  of the breath chamber  30  in between the rear frit plate  26  and the breath exit hole  40 . The pressure measurement hole  42  is defined in the wall within the distal space  38  and is essentially a tap for measuring ambient and distal pressure differential. The pressure of the breath in distal space  38  via pressure measurement hole  42  pressurizes the existing air within the pressure channel (pressure path). The pressure path extends through the pressure measurement hole  42  into the channel  44  and is channeled over and then down through pressure tunnel  46  and to the handle assembly  12 . The pressure that is inside the distal space  38  of the breath chamber  30  is being measured by pressure transducer  50  and based on the pressure measurement the flow rate through the breath chamber  30  can be calculated. The electronics of the pressure transducer  50  are located in the handle assembly  12  (i.e., on the circuit board  108 ). 
     In use, once the analysis cartridge  14 , is placed on the handle assembly  12 , a user blows through the mouthpiece  18  and the breath chamber  30  so that a predetermined volume of air or breath (e.g., 3 liters) is exhaled through the breath chamber  30 . Therefore, CCM are filtered out of a predetermined or known volume of breath and are collected on the substrate  28 . After the CCM have been collected, the user removes the analysis cartridge  14  from the handle assembly  12 , removes the mouthpiece  18  and places the analysis cartridge  14  in the analysis device  16 , as described below. The analysis cartridge  14  and handle assembly  12  in combination (shown in  FIG. 6B ) are referred to herein as the breath capture assembly  13 . 
     As shown in  FIGS. 6A-7 , the analysis cartridge includes a bottom window and two side windows. The bottom window is referred to herein as the fluorescence window  170  and the side windows are referred to herein as the light entry window  172   a  and the light exit window  172   b . The fluorescence window  170 , light entry window  172   a  and light exit window  172   b  are used in conjunction with an optical system  77  (also referred to herein as a fluorescence detection assembly or fluorometer) in the analysis device  16  described below. In a preferred embodiment, the windows  170  and  172   a  and  172   b  are a clear plastic and are the same material as the remainder of the main body portion  11 . However, the windows can be a different material. Preferably, the windows  170  and  172   a  and  172   b  are optically polished and are oriented such that outer surface is orthogonal to the appropriate components of the optical system  77  (described below). In a preferred embodiment, the remainder of the main body portion  11  is not optically clear and includes a draft so that the windows  170  and  172   a  and  172   b  are isolated (i.e., the sides or bottom of the cartridge are angled or not parallel to the outer surface of the windows). Preferably, the light entry and light exit windows  172   a  and  172   b  are parallel to one another and the fluorescence window  170  is perpendicular to the light entry and light exit windows  172   a  and  172   b.    
     In a preferred embodiment, the analysis cartridge  14  is made of plastic (e.g., polycarbonate, PMMA, etc.) and the various pieces are ultrasonically bonded to one another. However, this is not a limitation and the analysis cartridge can be made of any desirable material and bonded as desired. 
       FIGS. 1 and 8-23  show the analysis device  16 . As shown in  FIGS. 1 and 8-11 , generally, the analysis device  16  includes a case  48  (comprised of two halves  48   a  and  48   b ), a door  54  that is slidable between open and closed positions, a shroud  56 , analysis pocket  58  defined in the shroud  56  (where the analysis cartridge is received), a bottom  57 , a main circuit board  74 , a rotation assembly  76  and a display  60  (which is preferably touch screen). It will be appreciated that the full analysis pocket  58  includes the tapering funnel portion  58   a  and the well  173  described below. The analysis device  16  also includes an on/off button  62 , a speaker  64 , a handle storage pocket  66 , a USB port  68  and a DC input power port  70  (see  FIG. 8 ). The analysis device  16  also includes a battery  72 , as shown in  FIG. 9  and a battery door  156  shown in  FIG. 11 . The battery is preferably rechargeable, however this is not a limitation. 
     Shroud  56  is the interface between the analysis cartridge  14  and the analysis device  16 . In use, after a breath sample is taken using the analysis cartridge  14 , the user places the analysis cartridge in the analysis pocket  58  (through pocket opening  58   b ) and closes the door  54 .  FIG. 10  shows the analysis device  16  with the bottom half of the case  48   b  removed. As shown, the analysis device  16  includes the main circuit board  74  and the rotation assembly  76  for mixing the elution solution  34  and aldehydes, as described below. The rotation assembly  76  also includes the optical system  77 . The main circuit board  74  (mother board) is the controller and includes (or is in electrical communication with), but is not limited to, the USB port  68 , DC input power port  70 , cable(s) for communicating with a motor  78  and the optical system  77  (and optics boards  160  and  162  described below), a cable for communicating with the handle assembly  12  (via plugs  107  and  148 , cable  106  and circuit board  108 ), the display  60 , on/of switch  62 , battery  72 , and optical sensors for sensing if the door  54  is open or closed. 
       FIG. 11  shows the analysis device  16  exploded and illustrates the first and second halves of the case  48   a  and  48   b , the slideable door  54 , the main circuit board  74  and the rotation assembly  76 . As shown, the analysis device  16  also includes an end cap  146  having a plug  148  therein for connection of the cable  106 , and a tube  150  that defines the handle storage pocket  66 . As discussed above, the handle storage pocket  66  and tube  150  include a magnet  150   a  that interacts with magnet  111  in the handle assembly  12 . The interaction of the two magnets helps hold the handle assembly  12  in the handle storage pocket  66 . In a preferred embodiment, the first half of the case  48   a  includes an opening  152  that aligns with the shroud  56  to define the analysis pocket  58 . Opening/cover  154  in the case  48  houses the display  60 . In a preferred embodiment, the second half of the case  48   b  includes the bottom  57 , battery door  156  and openings for the USB port  68  and DC input power port  70 , which are part of the main circuit board  74 . 
       FIGS. 12-20  show the rotation assembly  76  with most other components omitted. As shown in  FIGS. 12-13 , the rotation assembly  76  generally includes a rotatable portion  86 , first and second fixed members  88   a  and  88   b , motor  78 , a gear train  81  (that preferably includes a pinion gear  83  that drives a large gear  84 ), the shroud  56 , and the optical system  77 . Motor  78  includes a drive shaft (not shown) that drives pinion gear  83 , which meshes with and rotates large gear  84 . Motor  78  is preferably controlled by main circuit board  74 . It will be appreciated that the center portion (the rotatable portion  86 ) pivots, and the fixed members  88   a  and  88   b  stay stationary within the case  48 . Large gear  84  preferably includes an arcuate slot  186  therein that receives a guide protrusion  188  on fixed member  88   a . The ends of the arcuate slot  186  provide stops (by interacting with guide protrusion  188 ) so that the rotatable portion  86  can only rotate a certain degree in each direction. 
       FIGS. 16-17B  show the rotatable portion  86  alone. The rotatable portion  86  includes first and second axle members  164  and  166  that connect to the shroud via a key and keyway relationship. This is not a limitation. In another embodiment, the first and second axle members  164  and  166  can be glued or bonded to the shroud or can be unitary with the shroud. As shown in  FIG. 13 , in a preferred embodiment, the shroud  56  includes axially aligned cylindrical protrusions  174  with a key  56   a  on opposite sides thereof. The protrusions  174  receive first and second axle members  164  and  166  that include complementary keyways  176  defined therein. First and second axle members  164  and  166  are preferably keyed so that they only fit on the shroud  56  in one orientation. 
     The first and second axle members  164  and  166  have bearings  178  thereon that cooperate with central openings  180  in the fixed members  88   a  and  88   b  and allow the rotatable portion  86  to rotate. The rotatable portion  86  is connected to the large gear  84  via at least one key  166   a  that meshes with at least one keyway  84   a  in the center opening of the large gear  84 . The second axle member  166  also includes a stop  184  for the large gear  84  and a cable passing recess  165  that allows a cable (not shown) coming from the main circuit board  74  and extending to the optical system  77  to pass therethrough. Therefore, the motor  78  drives the pinion gear  83 , which drives the large gear  84 , which meshes with and drives the second axle member  166 , which drives the shroud  56  (which holds the analysis cartridge  14  in the analysis pocket  58 ) and all other components attached thereto, such as the optical system  77  and an arm  80  (discussed below). 
     In a preferred embodiment, the analysis device  16  includes a door lock assembly where the door  54  is locked by the motion of the rotation assembly  76 . Preferably, the door lock assembly includes a door detection sensor that senses whether the door is open or closed. The shroud  56  includes a cam feature  114  (see  FIG. 16 ) thereon that interacts with a pivotal tab member on the case. The cam feature  114  is positioned so that when the rotation assembly  76  goes to the load position the tab is disengage, thus allowing the door to slide open. In all the other orientations of the rotation assembly  76 , the tab is up and that locks the door and prevents it from sliding open. 
     As will be described below, the rotatable portion  86  rotates with the analysis cartridge  14  therein to mix the PD solution  35  therein and to allow the optical system  77  to perform its analysis. In addition, the rotatable portion  86  includes a cam and lever system to translate the rotational motion to pivotal motion, so that the arm  80  pushes the filter assembly  19  from the breath chamber  30  into the fluid chamber  32 .  FIGS. 17A-19C  show the cam and lever system and how the rotation assembly  76  moves the arm  80 . The arm  80  is pivotal on post  191  that defines a pivot axis and includes a first end  80   a  with a ball bearing  122  thereon that rides on a cam surface  120  and a second end that moves in and out of an arm opening  56   b  in the side of shroud  56 . In a preferred embodiment, the arm  80  is pivotal on post  191  and is secured to a tab  192  that extends from the first axle member  164 , as shown in  FIG. 17B . Preferably, the arm  80  includes an opening  189  that receives post  191 , which extends downwardly from the shroud  56  (see  FIG. 17B ). Spring  124  is formed so that the coil section forms an opening that receives post  191 . A fastener  190  extends through the opening  192   a  in tab  192  and into the end of post  191 . The first end  80   a  of the arm  80  with the ball bearing  122  extends into a cam channel  194  defined in second fixed member  88   b  (see  FIG. 18 ). 
     The arm  80  is movable between a stowed position ( FIG. 19A ) and a deployed position ( FIG. 19B ). Spring  124  biases the arm to the stowed position. As shown in  FIG. 18 , the cam surface  120  is curved and includes an increasing radius along the path that the ball bearing  122  travels. As the rotatable portion  86  (and arm  80 ) rotates, the ball bearing  122  rides on cam surface  120 . The increasing radius of the cam surface  120  causes the arm  80  to pivot about pivot axis and therefore push the second or working end  80   b  of the arm  80  into arm opening  56   b  in the shroud  56 . Compare  FIG. 19A  where the arm is not pivoted inwardly to  FIG. 19B  where it is pivoted inwardly. Due to the positioning of the analysis cartridge  14  within shroud  56 , the second end  80   b  of the arm  80  pushes the filter assembly  19  from the breath chamber  30  to the fluid chamber  32 . The cam surface  120 , ball bearing  122 , pivotal (and spring biased via spring  124 ) arm  80  work together to convert rotational motion into pivotal motion. 
       FIGS. 17B-17D and 20  show the optical system  77 . The optical system  77  includes a housing  196  comprised of first and second halves  197   a  and  197   b  (the second half  197   b  is omitted in  FIG. 20 ). The housing  196  is preferably secured to the shroud  56  by threaded fasteners  196   a  (see  FIG. 10 ). In a preferred embodiment, the shroud  56  includes four fastener receiver members  175  that receive the elongated threaded fasteners  196   a  that extend through complementary first and second receiver tubes  177   a  and  177   b  on the first and second housing halves  197   a  and  197   b . The fastener receiver members  175  can be internally threaded or can be made so that the threads are created in the plastic as the threaded fastener  196   a  is screwed therein. As shown in  FIG. 17B , an extra set of first and second receiver tubes  177   a  and  177   b  are included that do not correspond to a fastener receiver member  175  on the shroud  56 . The threaded fasteners  196   a  therefore secure the two halves of the housing together and secure the housing  196  to the shroud  56 . 
     Shroud  56  includes an analysis opening  200  in the bottom thereof through which the back of the analysis cartridge  14  extends when it is in the analysis pocket  58 . The analysis opening  200  is aligned with a well  173  in the second half  197   b  of the housing  196  that receives the back of the analysis cartridge  14  therein. The housing  196  includes an analysis cartridge receiving portion  204 , as shown in  FIG. 17C  that defines the well  173 . The housing  196  is formed such that the first and second housing halves  197   a  and  197   b  cooperate to define an light chamber  198 , a fluorescence chamber  207 , a light trap  94 , and the well  173 . The shroud  56  includes three recesses  203  on the bottom surface thereof that cooperate with recesses on the analysis cartridge receiving portion  204  to define a light entry aperture  216 , a fluorescence aperture  217  and a light trap opening  218 . 
     The optical system  77  also includes a first optics circuit board or microcontroller  162  that includes an LED  79  and a second optics circuit board  160  that includes a receiver or detector  82  (e.g., a photo diode). Both optics circuit boards include sockets or connectors  163  for connecting cables (not shown) for communication and control from the main circuit board  74 . The optical system  77  also includes at least a first lens  90  and at least a first filter  92  positioned in the light chamber  198 , and at least a second lens  96  and at least a second filter  98  that are positioned in the fluorescence chamber  207 . The housing  196  is formed such that the first and second housing halves  197   a  and  197   b  cooperate to define a first lens pocket  199 , a first filter pocket  201 , a second lens pocket  203  and a second filter pocket  205 . 
     As shown in  FIGS. 17C-17E and 20 , the top housing half  197   a  includes first and second recesses  212  and  214  defined therein that cooperate with first and second recesses  213  and  215  in the top surface of the analysis cartridge receiving portion  204  to at least partially form the light entry aperture  216  and the fluorescence aperture  217 . The top and bottom housing halves  197   a  and  197   b  also cooperate to at least partially form the light trap opening  218 . When the analysis cartridge  14  is positioned in the well  173 , the light entry window  172   a  is aligned with the light entry aperture  216 , the fluorescence window  170  is aligned with the fluorescence aperture  217 , and the light exit window  172   b  is aligned with the light trap opening  218 . 
     In use, the LED  79  shines light along a light path (LP) through the first lens  90 , through a first filter  92 , through light entry aperture  216 , through light entry window  172   a  in analysis cartridge  14  (where it causes the CCM in the fluorescing solution  206  to fluoresce within the sensing chamber  32   b ), through light exit window  172   b , through light trap opening  218  and into light trap  94 . The light trap  94  is configured with angled walls so that the light that enters therein bounces around and cannot escape back through the entry opening and be reflected in any way toward the detector  82 . The light reflected from the fluoresced CCM exits the analysis cartridge  14  along a fluorescence path (FP) through fluorescence window  170  at an approximately 90 degree angle from the light entering the analysis cartridge  14 . The fluorescence path travels through fluorescence aperture  217 , through the second lens  96  and the second filter  98  and to the detector  82 . This is generally an emitter detector set-up. In a preferred embodiment, the detector  82  is at about ninety degrees to the emitter  79 . Other angles are within the scope of the invention. 
     In a preferred embodiment, the light emitted from the LED  79  and directed along the light path LP is as collimated as possible. Preferably, the light chamber  198  is designed to eliminate as much light as possible that is not collimated. To accomplish this, the light chamber  198  includes at least the first lens  90 , and a series of baffles and apertures (described below) positioned in the light path LP.  FIG. 17D  shows the light path LP as being directed parallel to the axis of the light chamber  198 . However, some light emitted from the LED  79  may not extend parallel to the axis. See, for example, the dashed lines in  FIG. 17D . In a preferred embodiment the first lens  90  is a Fresnel lens. However, this is not a limitation on the present invention. In an exemplary embodiment, the first lens  90  is a Fresnel lens with a focal length of about 10 mm, overall dimensions of 25.4 mm×25.4 mm×22 mm with a lens diameter of 12.7 mm (the second lens can have the same properties). However, none of these dimensions are limiting. The first lens  90  is positioned and includes specifications such that it preferably focuses the light from the LED  79  inside of the sensing chamber  32   b , within the well  173 . In an exemplary embodiment, the collimated beam of light is approximately 4 mm in diameter in the center of the sensing chamber  32   b.    
     As shown in  FIG. 17C-17D , in a preferred embodiment, the light chamber  198  includes a first light baffle  244  positioned therein that includes a first light baffle aperture  244   a  defined therein. The first light baffle  244  is positioned along a light path LP before the first lens  90 . The light chamber  198  also includes a second light baffle  246  positioned therein that includes a second light baffle aperture  246   a  defined therein. The second light baffle  244  is positioned along the light path LP between the first lens  90  and the first filter  92 . Preferably, a third light baffle  248  that includes a third light baffle aperture  248   a  is positioned in the light path LP after the first filter  92 . Preferably, the first and second light baffles are orthogonal to the direction of the light path LP and the third light baffle is not orthogonal to the direction of the light path LP. It will be appreciated that the baffles and apertures are formed by the first housing half  197   a  cooperating with the second housing half  197   b .  FIG. 17D  only shows the second housing half  197   b.    
     After the light passes through the analysis cartridge passes through the light trap opening  218  and into the light trap  94 . In a preferred embodiment, the walls of the light trap are black, which will absorb the majority of light that enters.  FIG. 17D  shows a plan view of the light trap  94 . In a preferred embodiment, the light trap  94  includes curved walls that help disperse the light as it bounces off. However, as shown in  FIG. 17D , with respect to the light path LP, the walls form angles that are designed to absorb most light because they are black, but also to reflect any light that is reflected toward another wall so that virtually no light escapes back through the light trap opening  218 . The light trap  94  preferably includes a first wall  94   a  that receives the light after it enters the light trap. The first wall is preferably angled between about 25° and about 45° with respect to the light path LP. In an exemplary embodiment it is angled at  350  from the light path LP. The light trap  94  also preferably includes a second wall  94   b  that is angled from the light path LP. Preferably, it is not at a right angle with the light path LP. In an exemplary embodiment, as light enters the light trap  94 , every time it bounces off a different wall approximately 80% is absorbed and 20% is reflected. After bouncing off of a few walls with the 80% to 20% absorption to reflection ratio, the remaining light will be negligible. 
     The fluorescence path FP also includes baffles and apertures therein together with the second lens  96  and second filter  98 . As shown in  FIG. 17D , in a preferred embodiment, the fluorescence path FP includes therealong a first fluorescence baffle  250  and related aperture, which is the fluorescence aperture  217  that is formed by the second recess  215  in the top surface of the analysis cartridge receiving portion  204 , the second lens  96 , a second fluorescence baffle  252 , which includes a second fluorescence baffle aperture  252   a  defined therein and second filter  98 . Preferably, the second lens  96  has the same specifications as the first lens  90 . However, this is not a limitation and the two lenses can be different. In another embodiment either or both of the light path and the fluorescence path can include more than one lens therein. 
     It will be appreciated that the light emitted from the LED is not completely collimated. Therefore, the light baffles and apertures are provided to block some of the light that is reflected off of the inside of the light chamber  198  and other extraneous light. The apertures in the first, second and third light baffles  244 ,  246  and  248  have smaller diameters than the light chamber  198 , thereby causing the light baffles to block or eliminate non-collimated light and help create a more collimated beam traveling along the light path LP through the light chamber  198 . 
     In a preferred embodiment, the diameters of the first, second and third light baffles get smaller as they are encountered along the light path. In an exemplary embodiment, the light chamber  198  has an inner diameter of about 16 mm, the first light baffle aperture has an inner diameter of about 10 mm, the second light baffle aperture has an inner diameter of about 9 mm, and the third light baffle aperture has an inner diameter of about 5 mm. As for the fluorescence chamber  207 , in an exemplary embodiment, the first fluorescence baffle aperture has an inner diameter of about 5 mm and the second fluorescence baffle aperture has an inner diameter of about 7 mm. 
     As shown in  FIG. 17D , in a preferred embodiment, the first lens pocket  199 , first filter pocket  201 , second lens pocket  203  and second filter pocket  205  each include crush ridges therein that help maintain the lens or filter therein in a stable position and prevent it from vibrating. However, the crush ridges can be omitted. Also, the well  173  can include an alignment member  254  therein and a drain  256  for draining any fluid in the well  173 . 
     The first filter  92  is provided to filter unwanted wavelengths of light from the beam of light emitted from the LED  79 . Any filter is within the scope of the present invention. In a preferred embodiment, the first filter allows transmission of light in a first range. For example, the first filter can include a transmission region of 300 nm to 540 nm T&lt;0.0001%, OD&gt;6, a transition region 540 nm to 550 nm, 0%&lt;T&lt;100%, and a blocking region that blocks light between 550 nm and 800 nm, T&gt;90%. However, this is only exemplary and any filter is within the scope of the present invention. In a preferred embodiment, the second filter allows transmission of light within a second range. For example, the second filter can include a blocking region that blocks light between 300 nm and 555 nm. T&lt;0.0001%, OD&gt;6, a transition region 555 nm to 565 nm, 0%&lt;T&lt;100%, and a transmission region of 565 nm to 800 nm T&gt;90%. However, this is only exemplary and any filter is within the scope of the present invention. The second filter  98  is designed to block all LED light that somehow made it into the fluorescence chamber  207  and to only allow the fluorescent light through at a predetermined wavelength (e.g., a long pass filter). 
     In a preferred embodiment, to further separate the fluorescence signals, lock-in amplification is used. A lock-in amplifier is used to help eliminate signals that have an origin in background light (e.g., lights from the room, circuit boards inside the device, backlighting from the screen, and any other white source that could potentially reach the detector  82 ). In an exemplary embodiment of using this technique, the LED is blinked on and off at a first rate (e.g., between 400 HZ and 1000 HZ). This helps get away from DC, which helps the noise issues. Then, if while detecting a signal is detected that does not have the same frequency and is very close in phase to the frequency at which the LED is being driven, there is a likelihood that the light is coming from some other source (e.g., background light), so it is eliminated from the signal. Generally, the lock-in amplifier takes the signal, it averages the signal from when the LED is on and then it averages the signal from when the LED is off and it subtracts the two. This preferably results in a fluorescence signal with little noise. 
     As shown in  FIG. 17C , In a preferred embodiment, the upper housing half  197   a  includes a lip or flange  258  that extends downwardly and overlaps another lip or flange  260  extending upwardly from the lower housing half  197   b . The complementary flanges help block light for entering or exiting the light chamber  198  or the fluorescence chamber  207 . Preferably, the flanges are offset from one another so that they overlap. 
     As discussed herein, the components of the optical system  77  are tuned specifically for the chemistry of an analysis cartridge and for this particular PD derivative and the amount of fluorescence that is to be measured. The ninety degree angle allows the photo detector  82  to detect the emitted light from the fluoresced CCM. In other words, the detector  82  is not receiving any light from the LED, but is only detecting the particles of aldehyde that get fluoresced. 
     From the description herein, it should be understood that the breath analysis system is used preferably to obtain a breath sample from a patient in the analysis cartridge  14  and then the breath sample is analyzed in the analysis device  16 . Once the analysis cartridge  14  is placed in the analysis pocket  58  in the analysis device  16  and extends down into the well  173 , the rotation assembly  76  rotates the analysis cartridge  14  a number of times to mix the contents, to move the filter assembly  19  from the breath chamber  30  to the fluid chamber  32  and to let the optical system  77  perform an analysis. 
       FIGS. 21A-23  show the steps for how the rotation assembly  76  mixes the PD solution  35 , moves the filter assembly  19  from the breath chamber  30  to the fluid chamber  32 , mixes the PD solution and breath aldehydes (CCM) to form a fluorescing solution  206  and how the optical system  77  performs an analysis of the fluorescing solution  206 . Each of the figures shows a cross-sectional end elevation view of the rotation assembly  76  with the analysis cartridge  14  in the analysis pocket  58 . In a preferred embodiment, as shown in  FIG. 21A , when the analysis cartridge is placed in the analysis pocket  58  and the back extends through the analysis opening  200  down into the well  173 , an alignment member  208  is received in the breath exit hole  40 . 
     In  FIG. 21A , the analysis cartridge  14  is in the analysis pocket  58  in the rotation assembly  76  and is in the configuration as the clinician has just taken it off the handle assembly  12  and placed it in the analysis pocket  58  (referred to herein as the start position). In use, the rotation assembly  76  rotates the analysis cartridge  14  through at least one pivot or rotation to introduce or mix the PD derivative  24  into the elution solution  34 .  FIG. 21B  shows the orientation of the rotatable portion  86  and the analysis cartridge  14  in a second position after rotation (referred to herein as a first mixing position). The rotatable portion  86  can move between the start position and the first mixing position a predetermined number of times for proper stirring or mixing. As discussed above, the rotatable portion  86  is moved via motor  78  (see  FIG. 15 ), which rotates the rotatable portion  86  and basically sloshes the PD solution  35  back and forth to make sure that the PD derivative  24  is completely in solution. This is the mixing step. 
       FIG. 21C  shows the orientation of the rotatable portion  86  and the analysis cartridge  14  in a third position (referred to herein as a baseline reading position). At this step (referred to herein as a baseline reading step), the analysis cartridge  14  is pivoted so that the fluid chamber  32  is straight up and down. Therefore, all of the PD solution  35  is in the rear portion or sensing chamber  32   b  of the fluid chamber  32 . At this point a baseline fluorescence reading is taken by the optical system  77 . In a preferred embodiment, this is done by turning on the LED  79  (the main circuit board  74  communicates with the first optical circuit board  162 ) and measuring the fluorescence of the PD solution  35  without any CCM therein, using detector  82 . The baseline reading is communicated by the second optical circuit board  160  to the main circuit board  74 . 
     Next, as shown in  FIGS. 21D-21E , the rotatable portion  86  and shroud  56  (and the analysis cartridge) move through and past the first mixing position (shown in  FIG. 21D ) and to a fourth position shown in  FIG. 21E  (referred to herein as the insertion position), where the arm  80  pushes on the filter assembly  19  and moves it along the filter assembly path P 2  (see  FIG. 2 ) from the breath chamber  30  to the fluid chamber  32 . In other words, in this step, the filter assembly  19  is inserted into the fluid chamber  32  by the arm  80 . When this happens, the second breakable barrier  36   b  is broken. It will be appreciated that the arm  80  pushes filter assembly  19  as a result of the cam path  120  discussed above. As shown in  FIG. 21D , the arm  80  is still in the stowed position after the baseline reading step. Therefore, during the mixing step and the baseline reading step and the rotation between the start position, the first mixing position, and the analysis position, the cam path  120  is configured such that the arm  80  remains in the stowed position. However, when the rotatable portion  86  rotates beyond the first mixing position, the increasing radius of the cam surface  120  pushes the ball bearing  122  outwardly, thereby pivoting the second end  80   b  of the arm  80  and pushing the filter assembly  19  inwardly, as shown in  FIG. 21E . In this position, all of the fluid is down in the front portion  32   a  of the fluid chamber  32  and the filter assembly  19  (frit plates  26  and substrate  28 ) is now in the fluid chamber  32 . However, the PD solution  35  has not yet touched any of the frit plates  26  or substrate  28  because of the fluid volume. 
     Next, as shown in  FIG. 22 , the rotatable portion  86  and shroud  56  (and the analysis cartridge) rotate to a fifth position (referred to herein as the analysis position), where the fluid chamber  32  is once again straight up and down. It will be appreciated that the positioning of the rotatable portion  86  is the same in the analysis position and the baseline reading position. In this position, the PD solution  35  filters through opening  17  in the frit stack holder  20  and the frit plates  26  and the substrate  28  thereby immersing the frit plates  26  and the silica  28  in the PD solution  35 . Also, the arm  80  has retracted back to the stowed position, but the filter assembly  19  has stayed in place. As the PD solution drains down and drips through the frit plates  26  it washes the CCM off the substrate  28  and into solution (referred to herein as the fluorescing solution  206 ). The analysis cartridge  14  is left in this orientation for a predetermined amount of time; enough time for the PD solution  35  to drain through and collect in the sensing chamber  32   b  of the fluid chamber  32  (the drainage step). During this step, the CCM are labeled or painted with the PD solution. In another embodiment, another mixing step can be added to further mix the fluorescing solution. Next, a fluorescence reading is taken by the optical system  77  to analyze the fluorescing solution. The original reading was the baseline without any CCM in the solution and now a measurement with CCM is taken. 
     After the analysis step, the rotation assembly  76  rotates to a sixth position, which is the same as the first position. In other words, the rotatable portion  86  returns to the start position so that the analysis cartridge  14  can be removed, as shown in  FIG. 23 . The analysis cartridge  14  can then be disposed. In a preferred embodiment, all the steps described above are done automatically. Basically, the user opens the door  54 , puts the analysis cartridge  14  in, closes the door  54  and hits go or the like on the display  60 . 
       FIGS. 24-29  show another embodiment of the present invention where some of the steps discussed above with the analysis cartridge  14  are divided into system that includes a breath analysis cartridge  220  and a fluorescence analysis cartridge  222 . The two cartridges together are referred to herein as an analysis cartridge system  219 . The structure of both the breath analysis cartridge  220  and a fluorescence analysis cartridge  222  is similar to the analysis cartridge  14  described above so that they can fit into the analysis pocket  58 , as described below. Like numerals in  FIGS. 24-29  refer to like components in  FIGS. 1-23 . 
     The analysis cartridge system  219  is used to capture breath aldehydes (CCM) and analyze them with the optical system  77  in the device similar to the breath cartridge and system described above. The general steps in using the analysis cartridge system  219  are as follows: 1) blow through the breath chamber  30  in the breath analysis cartridge  220  to capture CCM; 2) place the breath analysis cartridge  220  in the analysis pocket  58 ; 3) allow the rotation assembly  76  and arm  80  to move the filter assembly  19  from the breath chamber  30  to the fluid chamber  32  where the CCM mix with the elution solution  34  to form a CCM solution; 4) remove the breath analysis cartridge  220  from the analysis pocket  58 ; 5) move the ampule member from the first position to the second position to allow the CCM solution to mix with the PD derivative to form painted CCM solution  209 ; 6) connect the breath analysis cartridge  220  to the fluorescence analysis cartridge  222  so the painted CCM solution drains into the upper chamber of the fluorescence analysis cartridge  222  and through the filter assembly  19 . The substrate  28  in the filter assembly  19  captures the painted CCM from the painted CCM solution  209  and the absorption member  238  absorbs the remaining solution; 7) place the fluorescence analysis cartridge  222  in the analysis pocket  58 ; 8) allow the rotation assembly  76  and arm  80  to move the filter assembly  19  from the breath chamber  30  to the fluid chamber  32  where the painted CCM is eluted into a second elution solution  202  to form the fluorescing solution  206 ; 9) perform a fluorescence detection analysis of the fluorescing solution  206  with the optical system  77 . In a preferred embodiment, the second elution solution rinse comprises greater than 50% acetonitrile and preferably 90% ethanol. 
     As shown in  FIG. 25 , the breath analysis cartridge  220  includes an upper or breath chamber  30 , a fluid chamber  32  and a filter assembly  19  (with substrate  28  therein). A cap  221  plugs and seals the fluid chamber  32 . Breath analysis cartridge  220  includes an ampule assembly  224  positioned in the back of the fluid chamber  32 . Elution solution  34  is disposed in the fluid chamber  32 , and the fluid chamber  32  is sealed from the breath chamber  30 . This can be done by a breakable foil barrier, as described above or by another scaling method. For example, the flit stack holder  20  can seal the filter assembly opening  134 . In use, a patient blows through breath chamber  30  so that breath aldehydes are collected on the substrate  28 . Next, the breath analysis cartridge  220  is placed in the analysis pocket  58  in the start position (see  FIG. 21A ). Then the rotation assembly  76  rotates the shroud  56  and breath analysis cartridge  220  to the insertion position (see  FIG. 21E ) so that the filter assembly  19  moves from the breath chamber  30  to the fluid chamber. In the fluid chamber the CCM are eluted into the elution solution  24  to form the CCM solution. The breath analysis cartridge  220  is then removed from the analysis pocket  58 . 
     In a preferred embodiment, the ampule assembly  224  includes an ampule member  226  that is received in and slidable within a slide tube  228 . The ampule member  226  is preferably a cylinder that defines an interior  226   a , includes enclosed ends and has at least one and preferably two fluid openings  230  defined in the sidewall thereof. The end that protrudes outside of the fluid chamber is enclosed with a breakable barrier  231 . As shown in  FIG. 25 , the PD derivative  24  is disposed in the ampule member interior or fluorescence chromophore space  226   a . The ampule member  226  is movable within the slide tube  228  between a first position where the PD derivative  24  are separated from the fluid chamber  32  and a second position where the ampule member interior  226   a  is in communication with the fluid chamber  32 . In a preferred embodiment, when the ampule member  226  is in the first position, the fluid openings  230  are positioned inside the slide tube  228  and are therefore sealed from allowing the elution solution  34  therein, as shown in  FIG. 25 . However, when the ampule member  226  is slid to the second position, the openings  230  are now in flow communication with the fluid chamber  32 , which allows fluid in the fluid chamber  32  to enter the ampule member interior  226   a .  FIG. 26  shows the ampule member  226  in the second position. As shown in  FIGS. 24-25 , in a preferred embodiment, the ampule assembly  224  is housed in a receiver member  232  that mates with the fluorescence analysis cartridge  222  as described below. In use, after removing the breath analysis cartridge  220  from the analysis pocket  58 , as described above, the user presses the ampule member  226  and moves it from the first position to the second position. 
     As shown in  FIG. 27 , in a preferred embodiment, the fluorescence analysis cartridge  222  includes an upper chamber  30 , a fluid chamber  32  and a filter assembly  19  (with substrate  28  therein). The second elution solution  202  is disposed in the fluid chamber  32 . A cap  221  plugs and seals the fluid chamber  32 . A piercing member  234  is disposed in the upper chamber  30  adjacent the front opening  33 . The piercing member  234  is a hollow tubular member that includes a main body portion  235  with a piercer  236  extending therefrom. The piercer  236  has a smaller diameter than the main body portion and the upper chamber  30 . 
     In use, as shown in  FIGS. 28-29 , the receiver member  232  of the breath analysis cartridge  220  is inserted into the front opening  33  of the fluorescence analysis cartridge  222 . The piercer  236  then pierces the breakable barrier  231  thereby communicating the ampule member interior  226   a  with the lower chamber  32  of the fluorescence analysis cartridge  222 . When the breakable barrier  231  is pierced, the painted CCM solution  209  flows into the upper chamber  30  of the fluorescence analysis cartridge  222  and washes over the filters  26  and substrate  28  in the filter assembly  19  and the painted CCM are captured by the substrate  28 . Any excess solution is absorbed in the absorption member  238  in the rear of the upper chamber  30 . 
     The fluorescence analysis cartridge  222  is then placed in the analysis pocket  58  in the analysis device  16  in the start position (see  FIG. 21A ). The rotation assembly  76  then rotates the fluorescence analysis cartridge  222  to the insertion position (see  FIG. 21E ) where the painted CCM is eluted into the second elution solution  202  to form the fluorescing solution  206 . The rotation assembly  76  then rotates the fluorescence analysis cartridge  222  to the analysis position (see  FIG. 22 ) and a fluorescence analysis of the fluorescing solution  206 , as described above, is performed. 
       FIG. 30  shows another embodiment of an analysis cartridge  240 . The structure of analysis cartridge  240  is similar to the analysis cartridge  14  described above so that they can fit into the analysis pocket  58 , as described below. Like numerals in  FIG. 30  refer to like components in  FIGS. 1-29 . As shown in  FIG. 30 , the analysis cartridge  240  includes an upper chamber or breath chamber  30 , a fluid chamber  32 , a filter assembly  19  (with filters  26  separated by a substrate space  27 ) and a vent cap  242  to seal the fluid chamber  32 . Substrate that is pre-loaded with the PD derivative (referred to herein as PD substrate) is disposed in substrate space  27  and elution solution  34  is disposed in the fluid chamber  32 . 
     In use, the analysis cartridge  240  is placed on the handle assembly  12 , a user blows through the mouthpiece  18  and the breath chamber  30  for a predetermined amount of time and at a predetermined pressure (or within a predetermined pressure range) until CCM are collected on the PD substrate  241 . After the CCM have been collected, the user removes the analysis cartridge  240  from the handle assembly  12 , removes the mouthpiece  18  and places the analysis cartridge  240  in the analysis device  16 . 
     At first, the analysis cartridge  240  is in the analysis pocket  58  in the start position (see  FIG. 21A ). In a preferred embodiment, no first mixing or baseline reading step is needed. However, in another embodiment, these steps can be included. The rotation assembly  76  then rotates the analysis cartridge  14  through and past the position shown in  FIG. 21D  and to the insertion position where the arm  80  pushes on the filter assembly  19  and moves it along the filter assembly path P 2  from the breath chamber  30  to the fluid chamber  32 . In other words, in this step, the filter assembly  19  is inserted into the fluid chamber  32  by the arm  80 . It will be appreciated that the arm  80  pushes filter assembly  19  as a result of the cam path  120  discussed above. When the rotatable portion  86  rotates to the insertion position the increasing radius of the cam surface  120  pushes the ball bearing  122  outwardly, thereby pivoting the second end  80   b  of the arm  80  and pushing the filter assembly  19  inwardly, as shown in  FIG. 21E . In this position, all of the fluid is down in the front portion  32   a  of the fluid chamber  32  and the filter assembly  19  is now in the fluid chamber  32 . However, the elution solution  34  has not yet touched any of the frit plates  26  or the PD substrate  241  because of the fluid volume. 
     Next, as shown in  FIG. 22 , the rotatable portion  86  and shroud  56  (and the analysis cartridge) rotate to the analysis position, where the fluid chamber  32  is once again straight up and down. In this position, the elution solution  34  filters through opening  17  in the frit stack holder  20  and the frit plates  26  and the PD substrate  241  thereby immersing the frit plates  26  and the PD substrate  241  in the elution solution  34 . As the elution solution  34  drains down and drips through the frit plates  26  it washes the painted CCM off the PD substrate  241  and into solution (referred to herein as the fluorescing solution  206 ). The analysis cartridge  240  is left in this orientation for a predetermined amount of time; enough time for the elution solution  34  to drain through and collect in the sensing chamber  32   b  of the fluid chamber  32  (the drainage step). In another embodiment, another mixing step can be added to further mix the fluorescing solution. Next, a fluorescence reading is taken by the optical system  77  to analyze the fluorescing solution. 
     After the analysis step, the rotation assembly  76  rotates back to the start position so that the analysis cartridge  240  can be removed, as shown in  FIG. 23 . The analysis cartridge  240  can then be disposed. 
     The analysis device  16  and the screen  60  thereof includes the ability to walk a patient and the practitioner through the steps necessary to perform a breath analysis. For example, the screen provides a user with feedback on, for example, the flow rate to let a patient know if they are blowing too hard or too soft. 
     An exemplary set of steps using the system  10  and the user interface (UI) on the screen  60  is described below. It will be understood that this is only exemplary and that steps can be rearranged and/or omitted or added as desired. Furthermore, it will be understood that all entries are being made on the UI. In another embodiment, the UI buttons and keypad can be manual buttons and a keypad. The practitioner steps are as follows: 1) Turn on the device by pressing the power button  62  located above the touchscreen. 2) Press the start button on the UI. 3) Press the list button on the bottom left of the UI keypad and select the practitioner name in the upper right corner. This auto-populates the Practitioner ID. Press the go button. 4) Enter the Patient ID in the Patient ID field (could be a number or the patient email address). Press the go button. 5) Enter the 6-digit lot number in the analysis cartridge lot number field. Press the go button. 6) Open the analysis cartridge package. 7) Remove the handle assembly  12  from the handle storage pocket  66 . Press the arrow button “&gt;” to continue. 8) Give the breath capture assembly  13  to the patient. Press the arrow button “&gt;” to continue. 9) Press start. 
     The patient steps are as follows: 1) Press “tap to start”. 2) Deliver breath sample by blowing through the mouthpiece  18 , keeping the circle in the green zone on the UI. A ring will appear on the outside of the circle that represents the total volume. Maintain the green circle until the ring shows 100% complete. 3) When 100% total volume is reached, stop blowing and give handle to practitioner. 
     Continued practitioner steps are as follows: 10) Press “next”. 11) Disconnect the analysis cartridge  14 ,  220  or  240  from the handle assembly  12 . Press the arrow button “&gt;” to continue. 12) Place the handle assembly back in the handle storage pocket  66 . Press the arrow button “&gt;” to continue. 13) Disconnect the mouthpiece  18  from the analysis cartridge  14 ,  220  or  240 . Press the arrow button “&gt;” to continue. 14) Open the door  54 . Press the arrow button “&gt;” to continue. Insert the analysis cartridge  14 ,  220  or  240  into the analysis pocket  58 . Press the arrow button “&gt;” to continue. 15) Close the door  54 . 16) Press “done”. 17) The analysis device  16  will begin processing the sample. When it is finished, it will display “100%.”  18 ) Tap to reveal the score. 19) Press “done”. 
     It will be appreciated that if the analysis cartridge system  219  is used, the final few steps will change. After step  16  the steps are as follows: 17) The analysis device  16  will push the filter assembly  19  in the breath analysis cartridge  220  to the second position. When it is finished, it will display “done.”  18 ) Open the door  54  and remove the breath analysis cartridge. 19) Press ampule member. 20) Connect breath analysis cartridge to fluorescence analysis cartridge to allow the painted CCM solution to enter fluorescence analysis cartridge. 21) Disconnect breath analysis cartridge from fluorescence analysis cartridge. 22) Insert the fluorescence analysis cartridge  222  into the analysis pocket  58 . Press the arrow button “&gt;” to continue. 23) Close the door  54 . 24) Press “done”. 25) The analysis device  16  will begin processing the sample. When it is finished, it will display “100%.” 26) Tap to reveal the score. 27) Press “done”. 
     If the device is connected to Wi-Fi, the device will automatically upload the test record to a portal. If not, it will store the score until it finds a secure connection. The score can also be uploaded via USB port  168 . 
     It will be appreciated that modifications to the invention can be made. For example, the mouthpiece can be non-removable. 
     The present invention is directed to a method and device useful for the detection, quantitation and assay of carbonyl containing moieties (“CCM”) including aldehydes, preferably in biological samples, and preferably at low concentrations in the biological sample. In this regard, CCM is defined to include one or more different carbonyl containing moieties. 
     As used herein, a “biological sample” is referred to in its broadest sense, and includes solid and liquid or any biological sample obtained from nature, including an individual, body fluid, cell line, tissue culture, or any other source. As indicated, biological samples include body fluids or gases, such as breath, blood, semen, lymph, sera, plasma, urine, synovial fluid, spinal fluid, sputum, pus, sweat, as well as liquid samples from the environment such as plant extracts, pond water and so on. Solid samples may include animal or plant body parts, including but not limited to hair, fingernail, leaves and so on. The preferred biological sample for one embodiment of the present invention is the breath of a human. 
     A CCM is a compound having at least one carbonyl group. A carbonyl group is the divalent group &gt;C=0, which occurs in a wide range of chemical compounds. The group consists of a carbon atom double bonded to an oxygen atom. The carbonyl functionality is seen most frequently in three major classes of organic compounds: aldehydes, ketones, and carboxylic acids. As used herein, “aldehyde” has its ordinary chemical meaning and the method of the present invention is useful in detecting the concentration of aldehydes in biological samples. In particular, the present invention is useful in detecting various forms of aldehydes including without limitation 1-hexanal, malondialdehyde, 4-hydroxynonenal, acetaldehyde, 1-propanal, 2-methylpropanal, 2,2-dimethylpropanal, 1-butanal, and 1-pentanal. 
     The amount of the CCM captured by the substrate may vary, but typically for a substrate consisting of 200 mg of 50-270 mesh (300-50 μm) particle with a bed diameter of 12.5 mm, generally, it will be equivalent to the amount in a human&#39;s breath after breathing into a tube like a breathalyzer. Preferably it will be from 75 to 0.1 ppb (400 to 4 pmoles) and more preferably from 20 ppb to 0.01 ppb (80 to 0.4 pmoles). 
     The invention is amenable to “mix &amp; read” and “real-time” assay formats for the detection of CCM. The invention can be applied to the detection of CCM in solution. The invention can be applied to the detection of trace CCM in the gas phase by the addition of a primary capture (on a substrate as discussed below) and release (elution from the loaded substrate as discussed below) process. Preferably in one step of the process, gas phase CCM, for example, aldehydes from the breath of a human, are captured on a substrate. 
     The substrate of the present invention is desirably formed from a solid, but not necessarily rigid, material. The solid substrate may be formed from any of a variety of materials, such as a film, paper, nonwoven web, knitted fabric, woven fabric, foam, glass, etc. For example, the materials used to form the solid substrate may include, but are not limited to, natural, synthetic, or naturally occurring materials that are synthetically modified, such as polysaccharides (e.g., cellulose materials such as paper and cellulose derivatives, such as cellulose acetate and nitrocellulose); polyether sulfone; polyethylene; nylon; polyvinylidene fluoride (PVDF); polyester; polypropylene; silica; inorganic materials, such as deactivated alumina, diatomaceous earth, MgSO 4 , or other inorganic finely divided material uniformly dispersed in a porous matrix, with polymers such as vinyl chloride, vinyl chloridepropylene copolymer, and vinyl chloride-vinyl acetate copolymer; cloth, both naturally occurring (e.g., cotton) and synthetic (e.g., nylon or rayon); porous gels, such as silica gel, agarose, dextran, and gelatin; polymeric films, such as polyacrylamide; and so forth. Preferably the substrate is a solid phase matrix of silica optionally spaced between frits. The size of the substrate is chosen so that a measurable amount of CCM is captured by the substrate. The size can vary but generally it is about 2 mL, preferably about 1 mL and more preferably about 0.25 mL. 
     The substrate typically consists of a bed of particles with 50-60 angstrom pores, with a 50-270 mesh (300-50 μm), and a mass of 75 to 300 mg, preferably 60-120 mesh (250-125 μm) with a mass of 100 to 200 mg and more preferably 50-120 mesh (210-125 μm) with a mass of 125 to 175 mg. 
     In another step of the process, a fluorescence chromophore such as a phenylene diamine derivative is added to an elution solution to form a phenylene diamine solution. Phenylene diamine derivatives useful in connection with the present invention include but are not limited to many phenylene diamine derivatives including without limitation meta-phenylene diamine (“mPDA”) and its derivatives, with mPDA preferred for detecting aldehydes including without limitation 1-hexanal. While certain p-PDA or o-PDA derivatives may be useful in the method of the present invention, they are not useful for detecting 1-hexanal as they yield a cloudy colloidal suspension which is not useful for the optical based detection discussed below. 
     Other phenylene diamine derivatives include the following or mixtures thereof: 
     
       
         
         
             
             
         
       
     
     where R 1 , R 2 , R 3 . R 4  are each independently selected from the group consisting of H, alkyl, substituted alkyl, alkoxy, substituted alkoxy, acyl, acylamino, acyloxy, amino, substituted amino, aminocarbonyl, aminothiocarbonyl, aminocarbonylamine, aminothiocarbonylamino, aminocarbonyloxy, aminosulfonyl, aminosulfonyloxy, aminosulfonylamino, amidino, carboxyl, carboxyl ester, (carboxylester) amino, (carboxyl ester) oxy, cyano, halo, hydroxy. SO3-, sulfonyl, substituted sulfonyl, sulfonyloxy, thioacyl, thiol, alkylthio, substituted alkylthio, acyl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocycles, and substituted heterocycles. 
     The mPDA derivative mPDA-orange (pyridinium,4-[2-[4-(diethylamino)phenyl-ethenyl]-1-[1-(3,5-diminobenzamide)-pentylamino-5-oxyhexyl]) leverages both a) the sensitivity to environmental changes and b) the potential to modulate the surfactant dependence of the mPDA-aldehyde induced polymerization. The scheme used to synthetize mPDA-orange is to conjugate mPDA to the styrylpyridinium moiety via an alkyl amide linker. 
     mPDA-orange exhibits a quantum yield increase as the molecule is incorporated into the aldehyde induced mPDA polymer. In addition, the excitation and emission properties of the styrylpyridinium moiety affords a FRET (Forster Energy Transfer) generated signal from the mPDA polymer. The styrylpyridinium moiety exhibits a broad excitation with a maximum at 470 nm and an emission maximum at 570 nm. The excitation profile provides sufficient overlap with the emission profile of the mPDA polymer to afford FRET based signal generation. A FRET based signal generation would be manifest by an excitation at the mPDA polymer (405 nm) and emission at the styrylpyridinium moiety emission at 570 nm. 
     A direct aldehyde induced polymerization of mPDA-orange alone does not generate a response signal due to quenching of the styrylpyridinium at the high concentrations required for induction of the polymer. A response would only be expected when the mPDA-orange is contained within a mixture of mPDA and mPDA-orange. Indeed, an aldehyde response is only observed when mPDA-orange is doped into mPDA at significantly dilute molar ratios mPDAimPDA-orange 1,000:1 to 10,000:1. An increase in mPDA-Orange emission at 570 nm is observed when excited at 405 nm when 1 μM hexanal is added to the system. The increase in emission is not observed when the mPDA-orange styrylpyridinium moiety is excited directly at 470-490 nm. The response is approximately 3× over the background, where the conditions are 7 mM mPDA, 5 μM mPDA-orange (molar ratio 15,000:1), 90 mM NaCl, 15% Ethanol, 0.1% SDS, 50 mM citrate at ph 2.5. The excitation is at 405 nm and the emission is at 575-585 nm. As can be seen, in the absence of aldehyde the background level remains fairly constant and auto induction leading to incorporation of mPDA-orange appears to be minimal. Though the response for mPDA-orange is much less  3 X versus  15 X for mPDA alone the derivative offers several advantages: 1) increase wavelength discrimination afforded by the largeStokes shift between excitation and emission and 2) the enhanced baseline stability. 
     In general, the concentration of the phenylene diamine derivative in the phenylene diamine solution ranges from 0.5 mM to 25 mM. For mPDA, the mPDA concentration in the phenylene diamine solution generally ranges from 0.5 to 21 mM, preferably from 2 to 10 mM, and optimally 5 mM for aldehydes such as 1-hexanal. Notwithstanding the foregoing, for mPDA-orange, it must be diluted into mPDA at a low molar ratio, preferably 1000-10,000. 
     In general, the elution solution includes a salt, a buffer, a surfactant, and an organic solvent. The concentration of the salt ranges can from 5 mM to 200 mM and preferably from 20 mM to 80 mM; the concentration of the buffer can range from 25 mM to 200 mM and preferably from 40 mM to 60 mM; and the concentration of the surfactant can range from 0.05% (1.7 mM) to 0.4% (13.9 mM), and preferably from 0.15% (5.2 mM) to 0.25% (8.7 mM). Optimally 0.2% or 6.96 mM is used. The salt can be any salt that does not negatively impact the fluorescing solution and controls salting effects in the elution solution, and may include NaCl, LiCl, KCl, sulfates and phosphates, and mixtures thereof, with NaCl preferred. 
     The buffer is employed to maintain the elution solution mildly acidic and preferably at a pH of between 2 and 4.5, more preferably 2.5. The buffer can be a borate buffer, a phosphate buffer, a citrate buffer, an organic buffer such as HEPES (1-piperazineethane sulphonic acid) or also a TRIS (tris(hydroxymethyl)aminoethane) buffer, preferably a citrate buffer for use in detecting aldehydes. 
     The surfactant can include sodium decyl sulfate, sodium dodecyl sulfate (“SDS”), sodium tetradecyl sulfate and Standapol ES-1, with SDS including the C10, C12 and C14 version of SDS is preferable. Trition X-100, Ninate 11, Georpon 71, Tetraonic 1357, Cremapor-e1, Chemal 1a-9, Silwet L7900, Surfynly468, Surfactant 10G, and Tween 80 might also be used but they did not provide good results with the preferred elution solution, the CCM 1-hexanal and mPDA. 
     In the absence of SDS the polymerization and aldehyde response as discussed below is severely inhibited. mPDA is highly water soluble and the presence of SDS may provide a scaffold for organizing and orientating mPDA into a matrix to facilitate the polymerization reaction. 
     The solvent can include an aqueous solution of EtOH, MeOH, propanol, and isopropanol, with 15% EtOH preferred. 
     The molar ratio of salt concentration to phenylene diamine concentration is important. Generally the ratio should range from 0.03 to 0.5. For the CCM 1-hexanal, a molar ratio of mPDA to NaCl of 0.165 was found to provide optimal response. 
     The temperature for practicing the method of the present invention preferably ranges from 15 to 35° C. with 25 to 30° C. more preferred. 
     For the aldehydes such as 1-hexanal, one preferred embodiment of the elution solution comprises 33 mM NaCl, 50 mM Citrate, pH 2.5, 15% EtOH, and 0.2% SDS. Other preferred elution solutions include 50 mM Citrate, pH2.5, 15% propanol and 0.4% sodium decyl sulfate. 
     Using the elution solution containing a phenylene diamine derivative, the CCM is eluted into the phenylene diamine solution to form a fluorescing solution. The CCM and the mPDA react to form a fluorogenic species, the presence of which in the fluorescing solution is detected by measuring the fluorescence emitted by the fluorogenic species in the fluorescing solution. 
     The aldehyde content is quantitated by monitoring the signal rise (end-point) and/or rate of signal change (kinetic) which varies as a function of aldehyde concentration for a given mPDA concentration, and comparing such data with a carbonyl population sample of the breath. In practice the impact of carbonyls other than the selected carbonyl must be filtered out. There are two general assay format or detection modes. They are generally described as end-point and kinetic. In an end-point assay the system is incubated for a set time and the signal read. The signal at that point reflects the amount of analyte in the system. For a positive assay, the greater the concentration of the analyte, the greater the signal increase. In a kinetic assay the rate of change is monitored for a set duration. The rate of change is correlated to the amount of analyte. Preferably the end-point assay is employed with the present invention. 
     Assay measurements can be made on a typical fluorescence spectrometer including conventional scanning spectrometer, plate-reader or LED/diode based spectrometer following standard assay practices. To illustrate, the data displayed in  FIG. 31  was acquired by mixing a total of 2 mL of the reaction solution and aldehyde into a standard fluorescence cuvette and measuring the intensity increase using an LED/diode spectrometer at particular time slices to simulate an end-point determination. The LED/diode spectrometer utilized consisted of an Ocean Optics Jazz spectrometer with LED source and diode detection coupled via fiber optics to a Qpod-e (Quantum Northwest) temperature controlled fluorescence sample holder. The 405 nm excitation was produced with a violet LED (volts: 3.3 V, I: 0.03 A). The signal was detected using a ILX-5118 diode detection with emission set at 495-505 nm band pass and 250 msec integration. Like most fluorescence based assays, optimal settings are dependent upon the throughput and stray light rejection characteristics of the spectrometer used and must be empirically determined for each instrument. 
     In one preferred embodiment, the phenylene diamine derivative reacts with the CCM in solution to produce a fluorescence emitting or fluorogenic species. It is believed that the phenylene diamine derivative oxidatively couples to the CCM and the phenylene diamine derivative polymerizes to dimers, trimers, oligomers and/or polymers. It is not clear if the CCM actually becomes part of the growing polymer, although the polymerization is modulated by the presence of CCM and there is a dose response. 
     The process of using a CCM to polymerize the phenylene diamine derivative may be described as dispersion polymerization. Poly-phenylene diamines have been used to construct nanostructures and colloidal dispersions of different shapes, tubes, spheres and the like. However, if the polymerization results in large high molecular weight structures then precipitation occurs in the solution, which, in the present invention, may handicap optical detection. Thus the ingredients used in the method of the present invention must be chosen to avoid having elements in the fluorescing solution that inhibit detection and quantitation of the CCM. 
     The present invention utilizes the ability of CCM to modulate (initiate, catalyze and accelerate) the oxidative coupling and polymerization of phenylene diamine derivatives to detect and quantitate trace aldehydes, ketones and carbonyl containing analytes in a biological sample. Oxidative coupling and polymerization of phenylene diamine generates chromophoric and fluorogenic species. In the case of mPDA and aldehydes, the formation of polymers or multimers gives rise to a broad optical absorbance band at 405 nm and an associated emission band at 505 nm. The monomer absorbance is found in the UV region &lt;350 nm. As a result the production of the polymer can be conveniently followed by either conventional absorbance or fluorescence spectroscopy. In this regard, it should be appreciated that the absorbance and emission bands may vary depending upon the CCM and phenylene diamine derivative chosen, but all such bands useful in practicing this invention are part of the invention. 
     For example, with reference to  FIG. 31 , the emission spectrum of the reaction of mPDA in the presence of 1 μM 1-hexanal as a function of time is shown. The conditions of the fluorescing solution are: 1 μM 1-hexanal, 5.4 mM mPDA, 33 mM NaCl, 50 mM citrate (pH 2.5), 15% EtOH, and 0.1% SDS. The emission increases dramatically as a function of time. 
     With reference to  FIG. 32 , the reaction and responses with and without aldehyde (“blank”) are observed. The conditions of the fluorescing solution are: 1 μM 1-hexanal, 5.4 mM mPDA, 33 mM NaCl, 50 mM citrate (pH 2.5), 15% EtOH, and 0.1% SDS. The extent of the emission increase and the rate of increase are dependent upon the concentration of aldehyde in the phenylene diamine solution. At greater aldehyde concentrations, a larger and more rapid signal increase is observed. In the absence of aldehyde, the “blank” under goes a slow gradual small signal rise indicative of the slow polymerization of mPDA under the conditions examined. The polymerization is presumably due to the presence of trace oxidants such as iron, reactive oxygen species and other initiators. With the addition of a CCM, a significant signal enhancement over the blank or background is observed. Of particular note is that the rate of change is easily followed. As a result the detection system is amenable to both kinetic and end-point assay designs and detection modalities. The response can be quantitated at specific time points, e.g., 15 minutes (time slice) or by monitoring the slope as a function of aldehyde. The kinetic rate is slow enough that rapid and high precision of reactant additions is not required. The modulation of the polymerization reaction by a CCM such as an aldehyde and its use as a CCM quantitative sensor is another novel discovery and application described in this specification. Other alternatives including include labeling, painting or tagging the CCM for subsequent analysis. 
     With reference to  FIGS. 33A, 33B and 33C , the CCM induced polymerization reaction with the phenylene diamine derivative is shown to be sensitive to environmental conditions, and components of the reaction system such as the concentration of SDS. The conditions of the fluorescing solution in these figures are: 1 μM 1-hexanal, 5.4 mM mPDA, 33 mM NaCl, 50 mM citrate (pH 2.5), and 15% EtOH. For example, the reaction and aldehyde assay performance is dependent upon salt content, mPDA content, surfactant, pH and temperature. Since the reaction involves a “quasi-phase” transition from monomer to polymer insufficient mPDA concentration yields a slow reaction with limited signal change. In contrast, a large excess of mPDA results in a very rapid reaction and the formation of insoluble precipitates that limit optical detection. In addition, a large excess results in increased background or “blank” signal. 
     With reference to  FIG. 33A , the signal increases as function of SDS concentration. At an SDS concentration of 0.4%, the signal increase is almost 3 times the signal observed at 0.2%. 
       FIGS. 33B and 33C  show a comparison of the aldehyde response versus the blank for 0.2% SDS and 0.4% SDS, respectively. The increase in SDS concentration also results in an increase in “blank” or background signal. Both the signal and background are modulated by SDS concentration and the optimized SDS concentration cannot be determined by monitoring the signal response alone. As a result the SDS concentration must be optimized to provide the greatest discrimination between signal and background signal generation. For the embodiment specified, the optimal SDS concentration falls within a narrow concentration band, and small deviations can result in increased variability and limit the assay sensitivity. 
     With reference to  FIG. 34 , the fluorescence response for mPDA as a function of 1-hexanal concentration is displayed, with the background corrected. A linear response is observed from 0.1 to 1 μM 1-hexanal. The data points are the average of triplicate samples. The signal is measured at 20 minutes after the aldehyde is added to the phenylene diamine solution. Under these conditions, 10.8 mM mPDA, 65.5 mM NaCl, 50 mM citrate (pH 2.5), 0.2% SDS at 25° C., a solution limit of detection (LOD) of 0.1 μM can be achieved. 
     With reference to the chart in  FIG. 35 , mPDA exhibits a differential response for aliphatic aldehydes as a function of chain length. The chart reflects the fluorescence signal at 20 minutes after aldehyde addition, and the following conditions: 5.4 mM mPDA, 33 mM NaCl, 50 mM citrate (pH 2.5), 15% EtOH, and 0.1% SDS. The signal is measured at 20 minutes and this time-slice serves as pseudo end-point analysis method. For aliphatic aldehydes the relative response increases with aliphatic chain length. The response of acetylaldehyde is only 12% of the response observed for 1-hexanal. In contrast, the response of decyl (C 10 ) aldehyde is 30% greater than for 1-hexanal. 
     The nature of the aromatic diamine is also important to consider in employing the method of the present invention. O-PDA is highly reactive and undergoes rapid general oxidation. The high reactivity of o-PDA precludes its use as an aldehyde sensor in the preferred embodiment of the present invention. With reference to  FIG. 36 , the relative fluorescence response of a subset of diamines is displayed and illustrates the influence of both position and electronic effects on the aldehyde fluorescence response. Traditional aromatic electron donating and withdrawing effects should modulate the reactivity and susceptibility of the phenylene diamine derivative toward polymerization. An aldehyde response was not observed for both nitrophenylenediamine and naphthalenediamine under the preferred conditions, even when exposed to excess aldehyde. It has been found that aldehyde detection is based on the modulation of the polymerization of the reaction. If the molecule chosen is highly reactive and easily induced to polymerization then general oxidants can stimulate the reaction process and may limit its utility as a sensor. On the other hand, if the molecule is “too” stabilized, the polymerization process becomes inhibited and cannot be adequately stimulated by aldehyde and will require a much stronger oxidant to yield a response. 
     The present invention discussed above also includes a device for employing the method of the present invention. The device comprises a breath chamber preferably made of plastic and has a substrate in the breath chamber. The substrate is made from the materials discussed above and preferably silica. The substrate supports a carbonyl containing moiety from an animal&#39;s breath, e.g. aldehydes. The device also includes a fluid chamber. The fluid chamber includes an aqueous solution comprising an alcohol (e.g., 15% EtOH), a salt (e.g., NaCl), a surfactant (e.g., SDS), and a buffer (e.g. citrate). The solution can also comprise a phenylene diamine derivative such as mPDA. 
     The following example demonstrates one way to use the present invention to determine whether the sample breath of a human contains measurable aldehyde concentration and the concentration of the aldehyde in the breath. Employing the methodology discussed above, a series of fluorescence measurements are preformed to provide standards for various specific aldehydes and mixtures thereof that are known to be contained in a human breath sample (a population), and standards for concentrations of such various standards and mixtures thereof. Using these standards, the presence in a sample of human breath of a particular aldehyde or mixture of aldehydes and the concentration of such particular aldehyde or mixture of aldehydes can be determined. In general in one embodiment, the steps are as follows:
         a. Capturing the aldehydes from the human breath sample on silica;   b. Forming a solution comprising a salt, a buffer, a surfactant in an alcohol in mildly acidic conditions;   c. Adding a phenylene diamine derivative to the solution of step b;   d. Eluting the captured aldehydes into the solution of step c;   e. Determining the fluorescence signal of the solution of step c;   f. Determining the fluorescence signal of the solution of step d:   g. Subtract the fluorescence signal from step e from the fluorescence signal from step f; and   h. Comparing the net resulting fluorescence signal from step g with standard fluorescence of known aldehydes (a calibration curve, i.e., a response to known concentrations via an assay) to determine the concentration of aldehydes in the fluorescing solution. Simply put, this is a comparison of “y” axis values to provide the “x” axis value, or alternatively, solve of x knowing y and the calibration function y=f(x).       

     In another embodiment of the present invention, the substrate can be pre-loaded with an active reactive capture agent which covalently attaches to the CCM (the “Agent”) including without limitation a fluorescent hydrazine or aminooxy compound. Some examples of aminooxy compounds are as follows: aminooxy 5(6) tetramethylrhodamine (aminooxy 5(6) TAMRA), with a single isomer of either 5 or 6 preferred; and aminooxy 5(6) carboxyfluorescein (aminooxy 5(6) FAM), with a single isomer of either 5 or 6 preferred, for example aminooxy-C5-5-FAM. Others include aminooxy 7-amino-3-acetyl-4 methylcourmarin-6-sulfonic acid; 5-aminoxy acetic acid rhodamine B; and dinitrophenylhydrazin. In the foregoing examples, the reactive group is specified without the linkage group, which would be well known to those of skill in the art. In addition to the foregoing, the hydrazine or hydrazide versions are included within the present invention. Preferably the Agent is somewhat polar. 
     For example, for a substrate consisting of 200 mg of 50-270 mesh (300-50 μm) particle with a bed diameter of 12.5 mm, the amount of the Agent can be from 5.5 mg to 0.1 mg, and preferably from 2.5 mg to 0.4 mg. 
     In yet another embodiment of the present invention, a two-solution methodology is used. After the substrate is loaded with the CCM, the CCM is eluted into a first elution solution or “rinse” solution comprising generally 30% ethanol and preferably 50 mM citrate, 30% ethanol at ph 2.5. The Agent is added to the rinse solution thereby resulting in painted CCM. This solution is then passed through another substrate, preferably a silica frit stack, to capture the painted CCM. The painted CCM is then eluted from the substrate with the painted CCM captured therein using a second elution solution or “rinse” solution comprising greater than 50% acetonitrile and preferably 90% ethanol. One of the benefits of this second embodiment is that a baseline reading is not necessary to remove noise. 
     Unless the context clearly requires otherwise, throughout the description and the claims, the words “comprise,” “comprising,” and the like are to be construed in an inclusive sense, as opposed to an exclusive or exhaustive sense; that is to say, in the sense of “including, but not limited to.” As used herein, the terms “connected,” “coupled,” or any variant thereof, means any connection or coupling, either direct or indirect, between two or more elements; the coupling of connection between the elements can be physical, logical, or a combination thereof. Additionally, the words “herein,” “above,” “below,” and words of similar import, when used in this application, shall refer to this application as a whole and not to any particular portions of this application. Where the context permits, words in the above Detailed Description of the Preferred Embodiments using the singular or plural number may also include the plural or singular number respectively. The word “or” in reference to a list of two or more items, covers all of the following interpretations of the word: any of the items in the list, all of the items in the list, and any combination of the items in the list. 
     The above-detailed description of embodiments of the disclosure is not intended to be exhaustive or to limit the teachings to the precise form disclosed above. While specific embodiments of and examples for the disclosure are described above for illustrative purposes, various equivalent modifications are possible within the scope of the disclosure, as those skilled in the relevant art will recognize. Further any specific numbers noted herein are only examples: alternative implementations may employ differing values or ranges. 
     The above-detailed description of embodiments of the disclosure is not intended to be exhaustive or to limit the teachings to the precise form disclosed above. While specific embodiments of and examples for the disclosure are described above for illustrative purposes, various equivalent modifications are possible within the scope of the disclosure, as those skilled in the relevant art will recognize. Further, any specific numbers noted herein are only examples: alternative implementations may employ differing values, measurements or ranges. It will be appreciated that any dimensions given herein are only exemplary and that none of the dimensions or descriptions are limiting on the present invention. 
     The teachings of the disclosure provided herein can be applied to other systems, not necessarily the system described above. The elements and acts of the various embodiments described above can be combined to provide further embodiments. 
     Any patents and applications and other references noted above, including any that may be listed in accompanying filing papers, are incorporated herein by reference in their entirety. Aspects of the disclosure can be modified, if necessary, to employ the systems, functions, and concepts of the various references described above to provide yet further embodiments of the disclosure. 
     These and other changes can be made to the disclosure in light of the above Detailed Description of the Preferred Embodiments. While the above description describes certain embodiments of the disclosure, and describes the best mode contemplated, no matter how detailed the above appears in text, the teachings can be practiced in many ways. Details of the system may vary considerably in its implementation details, while still being encompassed by the subject matter disclosed herein. As noted above, particular terminology used when describing certain features or aspects of the disclosure should not be taken to imply that the terminology is being redefined herein to be restricted to any specific characteristics, features or aspects of the disclosure with which that terminology is associated. In general, the terms used in the following claims should not be construed to limit the disclosures to the specific embodiments disclosed in the specification unless the above Detailed Description of the Preferred Embodiments section explicitly defines such terms. Accordingly, the actual scope of the disclosure encompasses not only the disclosed embodiments, but also all equivalent ways of practicing or implementing the disclosure under the claims. 
     Accordingly, although exemplary embodiments of the invention have been shown and described, it is to be understood that all the terms used herein are descriptive rather than limiting, and that many changes, modifications, and substitutions may be made by one having ordinary skill in the art without departing from the spirit and scope of the invention.