Abstract:
A combination air sampling cassette and nutrient media dish having base, orifice plate, and nutrient media dish assembly for the collection of airborne particles. The orifice plate includes a plurality of holes that brings the nutrient media dish into fluid communication with the ambient air. A pump is connected to the air outlet in the base to pull air into the orifice plate through the holes, over the culture media, and out through the air outlet. As the air passes through the holes in the orifice plate it is accelerated and results in the selected impaction of particles in the culture media. A cover fits over the assembly to protect the culture media prior to and after sampling.

Description:
CROSS REFERENCES TO RELATED APPLICATIONS 
     This application claims the benefit of the filing date of U.S. Provisional Patent Application, Serial No. 60/163,012, filed Nov. 1, 1999, and entitled, Culturable Cassette. 
    
    
     BACKGROUND OF THE INVENTION 
     1. Field of the Invention 
     This invention relates generally to an improvement in the field of culturable air sampling for biological materials, and more particularly to a combination air sampling cassette and nutrient media dish for the selective collection of viable microorganisms from ambient air for culturing and analysis. 
     2. Discussion of Related Art 
     The growing awareness of the potential adverse health effects of microorganisms has given rise to an increased need to detect and quantify airborne microorganisms during evaluation of indoor air quality. As is well-known, many respirable viable particulates are entrained in the air we routinely breathe. The prior art includes a number of ambient air samplers adapted for use in collecting such viable microorganisms. In operation, these devices generally enable the collection of viable microorganisms onto a culture media or nutrient to stimulate incubation and foster colony growth. Subsequent laboratory analysis will identify and enumerate the colonies. 
     A well-respected device in wide use in the field of viable sampling is the Andersen viable (microbial) sampler, disclosed in U.S. Pat. No. 3,001,914. The patent teaches a mechanism which serves to count and classify microorganisms in air. The device comprises a series of stages, each stage including a perforated member positioned over a layer of nutrient media. Airborne particles are impacted onto the nutrient when air is drawn through the device. On incubation the viable particles in the nutrient become visible as colonies. The device is available as a six-stage or two-stage system when particle sizing is required, or single-stage when particle sizing is not required. 
     The Andersen sampler has two fundamental disadvantages: it is expensive and it is inconvenient to use. Several commercially available devices have been developed to overcome the limitations of the Andersen sampler, including, for example, the RCS™ centrifugal air sampler, the PBI surface air system sampler (SAS), and the Mattson-Garvin Slit-to-Agar air sampler. (RCS is a registered trademark of Biotest AG Corporation, Federal Republic of Germany.) While these devices arguably provide greater ease of use, they are still expensive and, more significantly, do not exhibit the collection efficiency of the Andersen sampler [Reference:“Evaluation of Eight Bioaerosol Samplers Challenged with Aerosols of Free Bacteria”, American Industrial Hygiene Association Journal (53), October 1992.] 
     Other devices similar to the Andersen design have been developed, including U.S. Pat. No. 3,922,905 and 4,038,057, both to Roth. The &#39;057 patent teaches a sampling device for removing particulate matter from gaseous media by jet impaction. The sampler includes a base, and impaction stage, and a connector diffuser connected to the impaction stage. The impaction stage has a plate with apertures for generating a prescribed velocity of the gas as it passes through the apertures. A nutrient medium plate is positioned under the apertured plate and the gas passing through the apertures impacts the nutrient medium such that airborne particulates are captured. The Roth devices were intended to have lower equipment cost and be simpler to use, but they have not displaced the Andersen sampler as the device of choice, most probably because they are not, in fact, appreciably easier to use than the Andersen sampler. 
     Accordingly, there remains a need for a viable sampler that has demonstrable equivalence to the Andersen sampler in terms of performance but which has the advantages of ease of use and lower cost. 
     Objects and Advantages 
     Accordingly, the primary objects and advantages of the combination air sampling cassette and nutrient media dish of the present invention include: 
     1. to provide a viable sampler that has lower equipment cost; 
     2. to provide a viable sampler that is faster and easier to use; and 
     3. to provide a viable sampler that is smaller and lighter than currently known devices. 
     Further objects and advantages of the invention will become apparent from a consideration of the drawings and ensuing description. 
     SUMMARY OF THE INVENTION 
     The combination air sampling cassette and nutrient media dish of the present invention generally comprises a base and media dish assembly that defines an upwardly opening recess therein, an air outlet therefrom, and an enclosure retaining culture media for the collection of particles. In a first preferred embodiment, an orifice plate comprising a plate with a plurality of small holes fits onto the integrated media dish in a sealing arrangement. A pump is connected to the air outlet in the base to pull air through the orifice plate, over the culture media, and out through the air outlet. As the air passes through the holes in the orifice plate it is accelerated and results in the selected impaction of particles in the culture media. A cover fits over the assembly to protect the culture media prior to and after sampling. 
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS 
     FIG. 1 is a cross-sectional side view in elevation of a first preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention, showing an ultra low profile configuration in which the orifice plate-media dish air passageway is substantially uniform in width; 
     FIG. 2 is a cross-sectional side view in elevation of a second preferred embodiment of the present invention, showing a low profile configuration of a combination air sampling cassette; 
     FIG. 3 is a top plan view of a variation of the inventive apparatus showing a variable width air passageway, said view showing the top cover and orifice plate removed; 
     FIG. 4 is a cross-sectional side view in elevation of a third preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention; 
     FIG. 5 is an exploded assembly perspective view of the low profile configuration of the cassette shown in FIG. 2, showing only the orifice plate, the media dish, the base, and the outlet seal; 
     FIG. 6 is a cross-sectional side view in elevation of a fourth preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention that utilizes a removable and reusable orifice plate with the air outlet contained in the combined base-media dish; 
     FIG. 7 is a cross-sectional side view in elevation of the cassette of FIG. 6 with the orifice plate removed and the cover in place; 
     FIG. 8 is an exploded isometric view of the cassette of FIG. 6; 
     FIG. 9 is a cross-sectional side view in elevation of a fifth preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention that utilizes a removable and reusable orifice plate with the air outlet contained in the orifice plate; 
     FIG. 10 is a cross-sectional side view in elevation of the air sampling cassette of FIG. 9 with the orifice plate removed and the cover in place; 
     FIG. 11 is an exploded isometric view of the cassette of FIG. 9; and 
     FIG. 12 is a perspective view of an orifice plate with a mounting flange and tapped hole. 
     FIG. 13 is a perspective view of a combination air sampling cassette and nutrient media dish with cut-outs in the media dish wall. 
    
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS 
     FIGS. 1 though  11  depict several different embodiments of the combination air sampling cassette and nutrient media dish of the present invention, generally denominated  10  herein. FIGS. 1 through 5 depict embodiments where the orifice plate  22  remains in place as an integral part of the air sampling cassette. FIGS. 6 through 11 depict embodiments of the design where the orifice plate  22  is removable and re-usable. 
     FIG. 1 is a cross-sectional side view in elevation of a first and most fundamental embodiment of the combination air sampling cassette and nutrient media dish  10  of the present invention. This embodiment is an ultra low profile configuration, comprising a base  42 , preferably substantially cylindrical and having a bottom  39 , a contiguous side  43  (preferably circumferential), and an integral contiguous interior partition  47  substantially concentric with said side  43  and thereby defining an integral culture media dish portion  45 . The cassette includes a removable cover  20 , and an orifice plate  22  interposed between the base  42  and the cover when the cover is on. The upper edge  41  of base side  43  extends slightly higher than the upper edge  49  of the media dish partition  47 , such that an air exit gap  62 ,  64  is defined by the orifice plate  22  and the upper edge  49  of the media dish  45 . The cassette further includes an outlet port  32  having an outlet seal  34  that may be selectively opened and closed. The side  43  of the base  42  and the partition  47  of the media dish  45  are spaced apart, preferably substantially evenly, to define an air passageway  40  of substantially uniform width  40 A and in fluid communication with the outlet port. Alternatively, as shown more clearly in FIG. 3, the width  40 B of the air passageway may be varied to channel air in desirable circulation patterns to facilitate impaction of microorganisms onto the nutrient media and to provide for a uniform and balanced flow of air through the orifice plate. This variation is applicable to each of the preferred embodiments. Thus, FIG. 3 is generic to each of the embodiments. A perimeter seal  38  between orifice plate  22  and the upper edge  41  of the side  43  prevents the introduction of atmospheric gases from entering the interior of the cassette without passing through the specifically sized apertures of the orifice plate. 
     In operation, a nutrient medium  30  is introduced into the media dish  42  and the cover is removed. A suction pump (not shown) is connected to the outlet port  32  to draw air through the orifices (see FIG.  5 ), and viable organisms and other particulate impact on the surface of the nutrient medium. The air then moves outwardly from the dish through air exit gap  56  and into air passageway  40  for ultimate exhausting through outlet port  32 . Both the exit gap and the air passageway may be varied in width. Also, the height of the media dish partition  47  may be varied over its length and may include a plurality of discrete cut-outs in the media dish partition  47  that may vary in size and distribution over its length, as shown in FIG.  13 . 
     FIG. 2 is a cross-sectional side view in elevation of a second preferred embodiment of the present invention, showing a low profile configuration of a air sampling cassette; and FIG. 5 is an exploded assembly perspective view thereof, showing the low profile configuration without a cover. In this embodiment, the air outlet port  32  opens at substantially the center  33  of base  36  and directly underneath the bottom  29  of the media dish  26 , which is supported above the interior bottom surface  27  of base  36  by media dish locating or support ribs  28 . From outlet port opening  35  an outlet duct extends to the discharge opening  32  for exhausting air. This configuration allows the media dish  26  to be removably placed within the base  36  and for air to flow substantially symmetrically around and underneath the media dish  26 , and then to exit from the side of the base  36  through the outlet port  32 . 
     FIG. 4 is a cross-sectional side view in elevation of a third preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention. This view shows a variation on the design of FIGS. 2 and 5, in which the outlet port  32  is underneath the base  36 , preferably though not necessarily at substantially the center of the base, and wherein base legs  44  elevate the base from the surface on which it is placed so that a pump connection can be made to the outlet port  32 . The advantage of this configuration is that it has a low tooling cost when the base  36  is molded. 
     FIG. 6 is a cross-sectional side view in elevation of a fourth preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention; FIG. 7 is a cross-sectional side view in elevation of the air sampling cassette of FIG. 6 with the orifice plate removed and the cover in place; and FIG. 8 is an exploded isometric view of the air sampling cassette of FIGS. 6 and 7. This embodiment utilizes a removable and reusable orifice plate  22  with the air outlet  32  integral with the combination base-media dish  42 . The orifice plate  22  includes a bayonet mounting means comprising a plurality of spaced-apart integral tangs  68  disposed downwardly from the underside  69  of the orifice plate and adapted for rotatable mating with complementary flanges  70  integral with the exterior circumference  78  of said base member  42 . As with the first preferred embodiment, shown in FIGS. 1 and 3, the media dish portion  45  is integral with the base and defined by a partition  47  interior to the side  43  of base  42 . 
     FIG. 9 is a cross-sectional side view in elevation of a fifth preferred embodiment of the combination air sampling cassette and nutrient media dish of the present invention. This design is a variation on the fourth preferred embodiment and also utilizes a removable and reusable orifice plate  22 . The distinction is that the air outlet  32  is integral with a contiguous circumferential side  57  of the orifice plate, rather than with the side of the base, and further that the bayonet mounting means involves the use of a plurality of interior slots  72  defining interior flanges integral with the base and adapted for mating with tangs  68  integral with the orifice plate  22 . FIG. 10 is a cross-sectional side view in elevation of the air sampling cassette of FIG. 9 with the orifice plate removed and the cover in place, and FIG. 11 is an exploded isometric view of the air sampling cassette of FIG.  9 . Each of the fourth and fifth embodiments involves the use of a gasket  38 , or O-ring, but may also employ any another method, to provide a secure seal between the orifice plate and base member. 
     As may be readily appreciated, the side  43  (FIGS. 1-7,  13 ),  78  (FIG.  8 ), and  57  (FIG. 11) of the cassette may either be integral with the base or integral with the orifice plate, and this variation determines the structure that defines the air passageway and the structure from which the air outlet port extends. When the side is integral with the orifice plate, as in FIG. 11, the upper edge  59  of the side is defined by the interior intersection of the interior top portion  61  of the orifice plate and the interior side  63  of side  57 . Whether the side is integral with the orifice plate or the base does not affect the principle of operation, and in either case the inventive combination obtains. 
     FIG. 12 is a perspective view of an orifice plate with a mounting flange  74  and a tapped hole  76 . 
     There are several advantages in having the media dish integral with the cassette base, as illustrated in FIGS. 1,  3 , and  6  through  11 . This allows for an extremely low profile design. At the preferred flow rate of 28.3 liters per minute the flow will be symmetrical as the air exits out of the integral media dish  45  and into the air passageway  40  before leaving through the outlet port  32 . However, at flow rates other than the preferred, the flow as the air exits the integral media dish  45  may not be symmetric. In these cases symmetry might be achieved by adjusting the width of the air passageway  40  as shown in FIG.  3 . In FIG. 3, the air passageway  40  varies in width as shown by the wide air passageway  48  and the narrow air passageway  50 . Another means of achieving a symmetrical flow would be to adjust the orifice plate-media dish air exit gap  56 , as can be seen in FIG.  1 . In FIG. 1, there is a large orifice plate-media dish air gap  62  on one side that is reduced to a small orifice plate-media dish air gap  64  on the opposite side. This change in gap size may be achieved by adjusting the design of the orifice plate  22  and or the media dish  45  (the media dish portion  45  is an integral part of the base  36  in this design configuration). Another means of achieving a symmetrical flow would be to vary the height of the media dish partition  47 , which may further include a plurality of discrete cut-outs  51  in the media dish partition  47  that may vary in size and distribution over its length, as shown in FIG.  13 . This feature is suitable for incorporation into any of the preferred embodiments of the inventive combination air sampling cassette and nutrient media dish. 
     The alternative embodiments provide for a removable media dish  26 , as illustrated in FIGS. 2,  4 , and  5 . A parameter of the base  36  in these embodiments is how the media dish  26  attaches and locates relative to the base  36 . Possible design configurations for this include, but are not limited to, the following: 
     1. Using locating ribs on the base  36  to locate the media dish  26  and using any method to hold it in place (solvent bonding, press fit, ultrasonic welding, spin welding, glue, snap into place, etc.). 
     2. Using ribs on the media dish  26  to center it within the base  36  and using any method of holding it in place (solvent bonding, press fit, ultrasonic welding, spin welding, glue, snap into place, etc.) 
     Culture media  30  is held in the media dish  26  or combined base-media dish  42 . Air impinges upon the media  30  during sampling. Viable organisms will then germinate and grow upon the media  30  for identification after sampling. It is important that the bottom of the media dish  26  is rigid so that it does not flex while the sample is being taken. 
     As is evident in the design variations of the above-described first through fourth embodiments of the present invention, possible variations of the media dish  26  would include, but are not limited to, the following: 
     1. Having stiffening ribs on the bottom (top side of the bottom or the bottom side of the bottom); 
     2. Having a post in the center of the plate, on the top surface, to support the bottom of the orifice plate  22 ; 
     3. Having the media dish  26  be an integral part of the base  36  (with or without the stiffening ribs or a center post); 
     The function of the orifice plate  22  is to accelerate and concentrate the air flow into discrete, countable, sections. It has a plurality of evenly spaced apertures (typically 200 to 400, with 400 being preferred) each having a small diameter, typically between 0.0100″ and 0.0465″ with 0.0100″ being preferred. The key to this part is the consistent diameter of the holes in the plate. The holes should be spaced roughly evenly from each other. In embodiments of the design where the orifice plate  22  is not removable and reusable it would be desirable, although not critical, that this part be transparent. Transparency aids in the analysis of the sampled part and in determining if a sample has been taken. 
     Several well-known means of manufacturing this part include, but are not limited to, the following: 
     1. Injection molding the holes into a unitary plastic part. 
     2. Stamping the holes into a piece of metal and insert molding the piece of metal into a piece of plastic. 
     3. Laser cutting the holes into a plastic, injection molded piece. 
     4. Drilling the holes into a plastic, injection molded piece. 
     5. Drilling the holes in a piece of metal and insert molding this into a piece of plastic. 
     6. Etching the holes into plastic. 
     7. Etching the holes into a piece of metal and insert molding this into a piece of plastic. 
     8. Machining the part from a solid block of metal and drilling the holes. 
     Another important function of the orifice plate  22  is to control the distance between the apertured surface of the orifice plate  46  and the media  30 . The design should be such that this distance is well controlled, will not vary due to manufacturing tolerances, and will not vary while the sampling is taking place. Consequently, the orifice plate  22  should not flex, nor should any of the supporting surfaces that hold it over the media  30 . Factors affecting this include the thickness of any walls or surfaces that are involved, including the plate with the holes, the use of stiffening ribs to minimize any flexure, and minimizing the distance and number of bends in getting from the holes to where the orifice plate  22  is supported. 
     The designs shown in the FIGS. 2,  4  and  5 , locate and keep the orifice plate  22  centered relative to the base  36  and to the media dish  26 ,  45  by using orifice plate locating ribs  24 . These ribs have lead-ins towards the outside that aid in placing the orifice plate  22  onto the assembly. Features of this configuration include: 
     1. The ribs  28  also hold the orifice plate  22  off of the media dish  26  a controlled amount, thus providing a high degree of control over the gap between the outlet of the holes in the orifice plate  22  and the top surface of the media  30 . The outer edge of the rim comes close to, but does not directly connect with the base  36  because that would reduce the amount of control over the gap between the outlet of the holes in the orifice plate  22  and the top surface of the media  30 . A seal is provided over the resulting gap between the orifice plate  22  and the base  36  by the perimeter seal  38 . 
     2. The outer edge of the orifice plate rim directly connects with the outer edge of the base  36 . This provides for a secondary seal in case the perimeter seal  38  were to fail. 
     3. Try to have both the outer edge of the orifice plate rim connect with the outer edge of the base  36  and have the orifice plate locating rib  24  connect with the edge of the media dish  26 . A means of facilitating this and allowing for manufacturing tolerances would be to have the orifice plate locating ribs  24  and or the media dish locating ribs  28  crush slightly, by design, and provide a snug fit. 
     4. Have the orifice plate locating ribs center and locate the orifice plate  22  relative to the media dish  26 . The bottom of the outer rim of the orifice plate  22  would engage with a gasket that also engages with the base  36 . This allows for larger manufacturing tolerances and controls the gap between the orifice plate  22  and the media  30 . 
     The function of the perimeter seal  38  is to prevent air from entering the cassette by any route other than through the orifice plate holes  46  and further provide attachment of the orifice plate  22  to the base  36  or combined base-media dish  42 . In the embodiments with a removable and reusable orifice plate  22 , the perimeter seal  38  is preferably a toroidal gasket or O-ring. FIGS. 8 and 11 show an O-ring that is retained within the orifice plate  22 . An alternative design would have a gasket applied to the combined base-media dish  42 . In such a case the method of attachment and of maintaining compression of the o-ring or gasket is by a twist-on connection. FIG. 8 shows a tang  68  on the orifice plate  22  that engages onto a flange  70  on the combined base-media dish  26 . FIG. 11 shows a tang  68  on the orifice plate  22  that engages into a slot  72  in the combined base-media dish  42 . Other attachment means including, without limitation, spring clamps, bolts and quick release nuts, and a hinging mechanism disposed at one end and an over-center clamp directly opposite. 
     Possible design configurations for embodiments with an integral orifice plate  22  include, among others, a piece of tape going around the perimeter of the assembly (around the joint that is formed between the outer edge of the rim of the orifice plate  22  and the upper, outer edge of the base  36 ); a snap together assembly with a small gasket to provide a seal; a bead of glue that has been applied on the joint; an ultrasonic weld; and a solvent bond. 
     Additionally, certain design configurations can show evidence of tampering with the sampling device. A piece of tape that has been torn, for example, would indicate that the cassette might have been opened. 
     In embodiments of the invention where the orifice plate  22  remains in place as an integral part of the air sampling cassette (FIGS. 1 through 5) the cover  20  and outlet seal  34  function to prevent air from getting into the cassette when it is not desirable to do so. 
     The cover  20  may be a very simple and low cost part. Possible design variants include, tape over the opening, a plastic cover that just rests on top of the rest of the assembly or is taped or otherwise held in place, a snap-on lid, a piece of parafilm, a sheet of plastic wrap, such as SARAN WRAP. 
     The outlet seal  34  is a very simple and low cost part. Possible design variants include, but are not limited to, tape, a plug that pushes into, over, or onto the outlet port  32  (as shown in FIG.  7 ), a snap-on piece that covers the outlet port  32 , a piece of Parafilm, and plastic wrap. 
     In embodiments of the invention where the orifice plate  22  is removable and reusable (FIGS. 6 through 11) the cover  20  has an integral cover seal  66  to prevent air from getting into the cassette when it is not desirable to do so. As shown in FIGS. 8 and 11, this is a gasket or O-ring. However, other design variants for the cover seal  66  include, without limitation, tape applied to a flange or other surface on the cover  20  and extending over a coplanar surface on the combined base-media dish  42 ; a piece of Parafilm; either of the foregoing methods applied to sealing on the outside wall of the combined base-media dish  42  in FIG.  8  and having an additional outlet seal  34  as described above; and any of the foregoing methods applied to sealing on the side  43  of the combined base-media dish  42  in FIG.  8  and having the outlet seal  34  sealed by a gasket from the inside and the gasket being affixed to the cover  20 . In embodiments of the design where the air outlet port  32  exits the base side  43 , an alternative design would be to recess the outlet port  32  into the base such that the outlet port  32  is below flush with the base side  43 . 
     A further embodiment of the invention would be a combined base-media dish  42  with an attached cover  20 . 
     FIG. 12 shows a orifice plate  22  with a mounting flange  74  and tapped hole  76 . These features are to enable the attachment of the orifice plate to a standard camera tripod stand by way of either screwing it directly to the stand or by the attachment of a quick disconnect mechanism. Another option for the mounting of the device to a tripod stand would be the provision of a cup being screwed to the tripod stand into which the assembled device would locate. 
     Operation of Invention 
     The overall purpose of the invention is to allow the user to take a culturable sample of the air. The procedure is different depending on whether the version of the device has a removable and reusable orifice plate  22  or not. For a device with an integral orifice plate  22  that remains in place the operation is as follows: 
     1. Cover  20  and outlet seal  34  are removed. 
     2. A pump is connected to the air outlet port  32  of the cassette. 
     3. The pump is turned on. 
     4. The pump is turned off and the time over which the sample was taken is recorded. 
     5. The cover  20  and outlet seal  34  are replaced. 
     For a device with a removable and reusable orifice plate  22  the operation is as follows: 
     1. Cover  20  (and outlet seal  34 , if applicable) is removed. 
     2. Orifice plate  22  is attached to the combined base-media dish  42 . 
     3. A pump is connected to the air outlet port  32 . 
     4. The pump is turned on. 
     5. The pump is turned off and the time over which the sample was taken is recorded. 
     6. Orifice plate  22  is removed. 
     7. Cover  20  (and outlet seal  34 , if applicable) is replaced. 
     In addition to knowing the amount of time that the pump was on, it is also important to know the flow rate of the air that was passing through the cassette while the sample was being taken. This flow rate is normally approximately 28.3 liters per minute. 
     As may be readily appreciated, the combination air sampling cassette and nutrient media dish of the present invention is a low cost, easy-to-use product that offers several advantages over the existing devices: It has lower equipment costs; it is lighter; it is smaller; and it is faster to use. The embodiments of the present invention having an integral orifice plate also have the advantages that they virtually eliminate the possibility for cross contamination between samples. It thus eliminates the need to clean any specialized equipment between samples. 
     While this invention has been described in connection with preferred embodiments thereof, it is obvious that modifications and changes therein may be made by those skilled in the art to which it pertains without departing from the spirit and scope of the invention. Accordingly, the scope of this invention is to be limited only by the appended claims.