Patent Description:
The present invention relates to a method and apparatus for testing of liquid samples and, more particularly, to a method and apparatus for determining the presence of at least two interferents in the liquid samples.

A wide variety of automated chemical analyzers are known in the art and are continually being improved to increase analytical menu and throughput, to reduce turnaround time, and to decrease requisite sample volumes. These clinical analyzers may conduct assays using reagents to identify analytes in a biological fluid sample such as urine, blood serum, plasma, cerebrospinal liquids, and the like. For convenience and safety reasons, these fluid samples are almost universally contained within capped sample containers (e.g., sample tubes). The assay reactions generate various signals that may be manipulated to determine a concentration of analyte in the sample. See, for example, <CIT> and <CIT> assigned to the assignee of the present application. Improvements in clinical analyzer technology have been accompanied by corresponding advances in pre- analytical sample preparation and handling operations such as sorting, batch preparation, centrifugation of sample tubes to separate sample constituents, cap removal to facilitate fluid access, and the like by automated pre-analytical sample preparation systems called Laboratory Automation Systems (LASs). LASs automatically transport sample in sample tubes to a number of pre-analytical sample processing stations that have been "linked together" like described in <CIT>and <CIT>.

These LASs may handle a number of different patient specimens contained in standard, bar code-labeled, and evacuated sample tubes. The bar code label may contain an accession number that may be coupled and correlated to demographic information that may be entered into a hospital's Laboratory Information System (LIS) along with test orders and other desired information. An operator may place the labeled sample containers (e.g., tubes) onto the LAS system, which may automatically sort and route the sample tubes to the requisite processing devices for pre-analytical operations such as centrifugation, decapping, and aliquot preparation prior to the specimen being subjected to clinical analysis by one or more analytical stations that may also be "linked" to the LAS.

For certain clinical assays, a serum or plasma portion (obtained from whole blood by centrifugation) may be used in the clinical analysis. To prevent clotting, an anticoagulant such as citrate or heparin may be added to the blood specimen immediately after it is originally obtained. Alternatively, the anticoagulant may be placed in an empty sample container (e.g., tube) prior to the patient sample being obtained. At a later time, the specimen may be centrifuged to separate the serum or plasma portion from the red blood cell portion. A serum separator may be added to the sample container to aid in the separation of the red blood cell portion from the serum or plasma portion.

After centrifuging and a subsequent de-capping process, the open sample container (e.g., tube) may be transported to an appropriate clinical analyzer that may extract liquid specimen from the sample container and combine the specimen with one or more reagents in special reaction containers (e.g., cuvettes or cups). Analytical measurements may then be performed, often using a beam of interrogating radiation interacting with the sample-reagent combination, for example, by using photometric or fluorometric absorption readings or the like. The measurements allow determination of end-point or rate values from which an amount of analyte related to the health of the patient may be determined using well-known calibration techniques. Unfortunately, the presence of certain components (e.g., colored interferents) in the sample as a result of some preexisting sample condition or processing may adversely affect the accuracy of the results of the analyte measurement obtained from the clinical analyzer.

In some cases, the integrity of the serum or plasma portion of the specimen may affect the interpretation of the results, i.e., the analyte reading of the clinical analyzer. For example, pre-analytical variables in the serum or plasma portion, which are not related to the patient disease state, may cause a different interpretation of the disease condition of the patient. Pre-analytical variables include hemolysis (ruptured red blood cells), icterus (excessive bilirubin), and lipemia (high, visible lipid content).

Typically, the integrity of the serum or plasma portion of the specimen is visually inspected by a skilled laboratory technician. This may involve a review of the color of the serum or plasma portion of the specimen. A normal serum or plasma portion has a light yellow to light amber color. Alternately, a serum or plasma portion containing hemolysis may be quite reddish in color, interferents may arise, for example, if an excess number of red blood cells are damaged, possibly during venipuncture, centrifugation, or after prolonged storage. When red blood cells are injured, they release low density, reddish-colored hemoglobin into the specimen causing a reddish-colored sample that is said to exhibit "hemolysis. " The presence of free hemoglobin (Hb) may be used to measure the degree of hemolysis and, when the hemoglobin concentration exceeds about <NUM>/dl, the hemoglobin may interfere with the colorimetric determination of analytes in the clinical analyzer due to the reddish interferent contained in the specimen.

A sample containing icterus may be dark yellow/brown in color. Such interferents may arise, for example, from an excess of bilirubin, the result of decaying red blood cells being converted in the spleen into bilirubin. Levels of bilirubin above <NUM>-<NUM>/dl are visibly yellowish and may, in particular, adversely affect enzyme-based immunoassays. Such a condition is termed bilirubinaemia or icterus.

A sample containing lipemia may be whitish in color. Interferents may arise, for example, as a whitish appearance in serum or plasma portion due to the presence of excess lipids. Such a condition is called lipemia and lipid levels above about <NUM>/dl may interfere with antibody binding in immunoassays and may accordingly also affect immunoassay results.

Thus, the degree of red color in a serum sample may correspond to the amount of hemolysis present in the serum or plasma portion of the specimen, the degree of dark yellow/brown color may correspond to the amount of icterus present in the serum or plasma portion of the specimen, and the degree of whitish color may correspond to the amount of lipemia present in the serum or plasma portion of the specimen.

Subsequent to centrifugation, when the red blood cell portion has been separated from the serum or plasma portion, a skilled technician may visually inspect the serum or plasma portion and, if judged to not have a normal light yellow to light amber color, the specimen may be discarded. Otherwise, the specimen will be processed and analyzed as ordered. However, visual inspection is very subjective, labor intensive, and fraught with the possibility of human error. Thus, various methods have been implemented to ascertain whether hemolysis, icterus, and lipemia (these three conditions are frequently called "HIL") are present in a serum or plasma portion of the specimen.

Typically, a laboratory technician will assign a hemolytic index, an icteric index, and a lipemic index to the serum and plasma portion of the specimen based upon the color. Based upon the value of the hemolytic index, the icteric index, and the lipemic index, the interpretation of the results from the clinical analyzer can be evaluated. Alternately, if the value of one or more of the hemolytic index, the icteric index, and the lipemic index are too high, the specimen may be discarded without analysis by the clinical analyzer.

As mentioned above, visual inspection can be labor intensive and costly. Furthermore, the possibility of human error exists with visual inspection, the results of the visual inspection may be highly subjective and may vary between workers, and one variable could mask or hide other variables. Furthermore, with closed container sampling, bar code labels directly on the container, and the use of automated clinical analyzers, the laboratory technician, in many instances, may simply not have a clear opportunity to visually observe the serum or plasma portion of the specimen. Thus, it is becoming increasingly important to evaluate the integrity of the serum or plasma portion of the specimen without the use of visual inspection by a laboratory technician.

One attempt to solve this problem involves optically viewing the serum or plasma portion of the specimen after the serum or plasma portion has been transferred to one of the cuvettes of the clinical analyzer. Measuring the optical characteristics of the specimen in the clinical analyzer eliminates the need for visual inspection. However, this test utilizes machine time of the clinical analyzer and, if the integrity of the specimen is determined to be compromised, additional machine time and a machine cycle are wasted. Furthermore, this procedure cannot be used with clinical analyzers that add reagents to the cuvette prior to adding the serum or plasma portion of the specimen.

<CIT> discloses monitoring a serum sample with a detector that performs a spectrophotometric analysis of the serum sample in the probe lumen through a substantially transparent section of the probe. From the spectrophotometric analysis, a hemolytic index, an icteric index, and a lipemic index of the serum sample can be established. Based upon these serum indices, the serum sample can be transferred to a clinical analyzer for additional tests or can be disposed of because the sample is compromised.

<CIT> discloses quality control material used to monitor instrument calibrations or used for recalibration for instruments that assess the amount of hemolysis, turbidity, bilirubinemia, and biliverdinemia, either separately, or any two, or any three, or all four digitizeously, in plasma or serum samples.

<CIT> discloses preliminarily testing a sample for HIL in the original incoming sample container, prior to being removed from the container and prior to being transferred to a clinical analyzer. In this approach, sample is not consumed and can be transferred to the clinical analyzer or a waste receptacle based upon results of the evaluation.

<CIT> discloses a method to reject a sample from further analysis based on determining the concentration of at least one interferent in the sample by: (<NUM> ) irradiating the sample with at least one frequency of radiation; (<NUM>) correlating absorbance of the radiation by the sample with a standard for the interferent(s) to determine the concentration of the interferent(s), and (<NUM>) rejecting the sample if the concentration of the interferent(s) exceeds a predetermined criteria.

One challenge in performing spectrophotometric analysis has been that the specimens are initially obtained in a variety of primary patient sample collection containers ("sample containers"). These sample containers are usually tubes of varying diameters and lengths. In the case of a patient blood sample, the liquid is often centrifuged to separate the liquid serum or plasma portion from the cellular phase (e.g., red blood cell portion). Such sample containers may have a patient identification label, varying and unpredictable amounts of the serum or plasma portion to be analyzed in the total specimen, and contain a relatively large amount of sample liquid.

Because of the problems encountered when endogenous interferents are contained within liquid samples to be clinically analyzed, there is an unmet need for a method and apparatus adapted to determine a presence of such interferents. The method and apparatus should not appreciably adversely affect the speed at which analytical test results are obtained and should allow making a determination on a relatively large sample portion so that the accuracy of such a determination is not affected. Furthermore, the method and apparatus should be able to be used even on labeled sample containers.

According to a first aspect of the invention, a method of determining the presence of at least two interferents in a clinical analysis specimen contained within a sample container according to claim <NUM> is provided.

According to another aspect of the invention, an apparatus adapted to determine the presence of at least two interferents in a clinical analysis specimen contained within a sample container according to claim <NUM> is provided.

The at least two interferents may be selected from a group consisting of lipemia, hemolysis, and icterus.

Still other aspects, features, and advantages of the present invention may be readily apparent from the following detailed description by illustrating a number of exemplary embodiments and implementations, including the best mode contemplated for carrying out the present invention. The drawings and descriptions are to be regarded as illustrative in nature, and not as restrictive. The drawings are not necessarily drawn to scale.

The following terms used in this application shall have the following meaning:
"Sample Tube" shall mean a blood collection tube used to collect blood from a patient. However, the sample tube may include any structural configuration adapted to receive and contain blood, such as tubular, slightly conical, etc. Additionally, the sample tube may be used for the initial collection of the blood from the patient or the blood may be transferred to the sample tube after being collected in another container. A suitable blood collection tube is manufactured by Becton Dickinson, located in Franklin Lakes, New Jersey.

"Serum Variables" shall mean and include hemolysis, icterus, lipemia, and other variables in the serum or plasma portion that may affect the accuracy of results of a clinical analyzer.

"Interferent" shall mean and include any of the serum variables, any disease condition, and/or any opacity, coloration, or particulate that may affect the interpretation of results of a clinical analyzer.

"Hemolytic index" shall mean the grade given to a particular sample based upon the estimated content of hemolysis present in the sample (specimen). Generally, the grading scale for visual observation ranges from zero through four (<NUM>-<NUM>). Zero represents substantially no hemolysis while four represents significant hemolysis. Alternately, the scale could be <NUM>-<NUM>, <NUM>-<NUM>, A-F, or some other range.

"Icteric index" shall mean the grade given to a particular sample based upon the estimated content of icterus present in the sample (specimen). Generally, the grading scale for visual observation ranges from zero through four (<NUM>-<NUM>). Similarly, zero represents substantially no icterus, while four represents significant presence of icterus. Alternately, the scale could be <NUM>-<NUM>, <NUM>-<NUM>, A-F, or some other range.

"Lipemic index" shall mean the grade given to a particular sample based upon the estimated content of lipemia present in the sample (specimen). Generally, the grading scale for visual observation ranges from zero through four (<NUM>-<NUM>). Similarly, zero represents substantially no lipemia, while four represents significant presence of lipemia. Alternately, the scale could be <NUM>-<NUM>, <NUM>-<NUM>, A-F, or some other range.

"Serum Indices" shall mean and include the hemolytic index, the icteric index, and the lipemic index.

"Predetermined Value" shall mean a value for the hemolytic index, the icteric index, or the lipemic index at which the integrity of the sample for testing may be considered to be compromised. The predetermined value varies according to the scale of the serum indices, which of the serum indices is in question, and the tests to be performed by the clinical analyzer or other device. For example, if the hemolytic index is rated on a scale of <NUM>-<NUM>, a hemolytic index of <NUM> could be considered to compromise the sample for some tests. Thus, the predetermined value in this example would be <NUM>. Alternately, a reading of <NUM> on a scale of <NUM>-<NUM> for the icteric index could be unacceptable in some instances. Thus, for this example, the predetermined value is <NUM>.

"Spectrophotometric analysis" shall mean and include measuring optical absorbance and/or reflectance, a turbidimetric analysis, a nephelometric analysis, and/or light scatter analysis at any angle or collection of angles. In general, the term "spectrophotometric" refers to capturing spectral response over a range of wavelengths and correlating a response for each of the wavelengths. A device that performs this analysis is referred to as a "spectrophotometer. " Such spectrophotometric analysis has been performed with near-infrared and adjacent visible radiation, which is capable of ascertaining hemoglobin, glucose, albumin, lipoproteins, and many other sera components.

The present invention provides a method and an apparatus for determining the presence of at least two interferents in a liquid specimen. The method may be carried out as a pre-analysis step prior to the liquid sample (specimen) being presented to a clinical analyzer for analytical analysis. In particular, one aspect of the present disclosure provides for delivering centrifuged samples for analytical analysis subsequent to being pre-inspected for a presence of at least two interferents like those that might be found within a blood sample. This aspect is accomplished by subjecting a liquid blood sample to an appropriate centrifugation to separate the sample into a red blood cell portion, and a blood serum or plasma portion. Subsequent to this centrifuging procedure, the blood serum or plasma portion of the sample (specimen) is tested for the presence of at least two interferents, such as hemolysis, icterus, and/or lipemia (hereinafter HIL) or other liquid non-uniformities therein (e.g., improper supernat level). If the sample is found to be free of interferents, it is allowed to continue through the system to be routinely processed for analytical analysis.

In one aspect, if the sample is found to contain more than a predefined amount of lipemia, then the sample may be rejected. The lipemic sample may then be subjected to a special pre-processing operation adapted to reduce an amount of lipemia therein. The specimen may then be allowed to be routinely processed for analysis or possibly retested for the presence of an interferent. In another aspect, if the specimen is found to contain more than a predefined amount of hemolysis, then the sample may be allowed to continue and be routinely processed for analytical analysis. However, the extent or degree of hemolysis may be reported along with the analytical results. Alternatively, the hemolyzed specimen may be subjected to a more sophisticated determination of the amount of hemolysis so that any analytical tests to be conducted on the specimen that are not affected by the presence of hemolysis may be routinely completed, and possibly a redraw of a fresh sample may be ordered and undertaken. If the specimen is found to contain more than a predefined amount of icterus, then the specimen may be allowed to be routinely processed for analytical analysis and the extent or degree of icteria may be reported along with the analytical results. The invention is based on the discovery that the extent of scatter or "bloom" of a collimated light beam directed through a centrifuged sample is a more rapid and less complex determination of a presence of an interferent (e.g., lipemia) in the specimen than other more sophisticated methods.

<FIG> (not part of the invention) shows an automated clinical chemistry sample handling system <NUM> (hereinafter "automated sample handling system") capable of automatically preprocessing multiple sample containers <NUM>, typically sample tubes (e.g., test tubes or blood collection tubes - see <FIG> (not part of the invention)), contained in multiple sample racks <NUM> prior to analysis by a clinical analyzer <NUM>, <NUM>, and/or <NUM>. However, any generally clear or transparent container may be used, such as a sample cup, cuvette, or other clear glass or plastic container. Typically, the clinical analysis specimens 20A (hereinafter "specimens") to be automatically processed may be provided to automated sample handling system <NUM> in the sample containers <NUM>, which may be capped with a cap 20B. Each of the sample containers <NUM> (e.g., sample tubes) may be provided with identification indicia and/or information 20C, such as a bar code, alphabetic, numeric, or alphanumeric indicia, that may be machine readable by one or more sensors <NUM>. The indicia and/or information 20C may indicate a patient's identification as well as the assay procedures to be accomplished upon the clinical analysis specimen 20A therein, for example. Such indicia and/or information 20C may be generally provided on a label 20D adhered to, or otherwise provided on the side of, the sample container <NUM>. Such labels 20D generally do not extend all the way around the sample container <NUM>. Accordingly, a window is provided along a side of the sample container <NUM> where the label 20D is not located and the clinical analysis specimen 20A may be viewed from the side in or through this window without interference by the label 20D. The sample containers <NUM> may be held in racks <NUM> that may have additional identification indicia thereon.

Automated sample handling system <NUM> may include an operating base <NUM> (e.g., a frame) upon which a conveyor track <NUM> (which may be belt-like) or other suitable conveyance mechanism or system transports individual sample containers <NUM> (e.g., sample tubes) carried in sample container carriers <NUM> from a sample container loading/unloading robotic station <NUM>, having one or more racks <NUM> as well as active input lanes, to a centrifuge <NUM> (e.g., an automated centrifuge). After being centrifuged, the sample containers <NUM> may continue on conveyor track <NUM> to a sample quality station <NUM> described hereinafter and adapted for automatically determining a presence of two or more interferents in the specimens 20A to be automatically processed by the automated sample handling system <NUM>. The specimens 20A may then be analyzed in the one or more analyzers <NUM>, <NUM>, and/or <NUM> before returning each sample container <NUM> (e.g., sample tube) to the sample container loading/unloading robotic station <NUM>. It should be understood that more than three analyzers <NUM>, <NUM>, and <NUM> may be linked by the conveyor track <NUM> but, for purposes of simplicity, only three are shown. Additionally, a remote analyzer <NUM> may be serviced by automated sample handling system <NUM> even though the remote analyzer <NUM> is not directly linked to the automated sample handling system <NUM>. For instance, an independent robotic system may carry specimens to the remote analyzer <NUM>. The automated sample handling system <NUM> may include a number of sensors (not shown) at one or more locations for detecting a location of sample containers <NUM> by means of reading identifying indicia or information (not shown) placed within each sample tube carrier <NUM>. In some embodiments, a distinct RFID chip may be embedded in each sample tube carrier <NUM> and conventional RFID reader systems may be employed in such tracking operations, for example.

Centrifuge <NUM> and each analyzer <NUM>, <NUM>, and <NUM> may be generally equipped with various robotic mechanisms <NUM> and <NUM>, <NUM> and <NUM>, or tracks <NUM> and <NUM>, respectively, for removing a sample tube carrier <NUM> from the track <NUM>, moving the sample tube carrier <NUM> to and from centrifuge <NUM>, to and from the analyzers <NUM>, <NUM>, and <NUM>, or facilitating movement of a sample container <NUM> into and out of the analyzers <NUM>, <NUM>, and <NUM>. Typically, the loading/unloading station <NUM> may include at least two X-Y-Z robotic arms <NUM> conventionally equipped with robotic clamping hands or fingers. However, any suitable robotic apparatus may be used.

Automated sample handling system <NUM> may be controlled by a conventionally-programmed computer <NUM>, preferably a microprocessor-based central processing unit CPU, which may be housed as part of, or separate from, the automated sample handling system <NUM>. The conventionally-programmed computer <NUM> may operate to control movement of the sample tube carriers <NUM> to and from the sample container loading/unloading robotic station <NUM>, the centrifuge <NUM>, quality control station <NUM>, and each clinical analyzer <NUM>, <NUM>, <NUM> (whereat various types of assay processing occurs) as described below. Computer <NUM> may control the automated sample handling system <NUM> according to software, firmware, or hardware commands or circuits such as those used on the Dimension® clinical chemistry analyzer sold by Siemens Healthcare Diagnostics Inc. of Deerfield, Illinois, and such control is typical to those skilled in the art of computer-based electromechanical control programming. However, any suitable electronic component or system for controlling the automated sample handling system <NUM> may be used.

The present invention may be implemented using a computer interface module (CIM) that allows for a user to easily and quickly access a variety of control screens and status information display screens. These screens may describe some or all aspects of a plurality of interrelated automated devices used for sample preparation and clinical analysis of a patient's specimen. Such a CIM preferably employs a first display screen that is directly linked to a plurality of additional display screens containing on-line information about the operational status of a plurality of interrelated automated devices as well as information describing the location of any specific sample and the status of clinical tests to be performed on the sample. The CIM is thus adapted to facilitate interactions between an operator and automated sample handling system <NUM> wherein the CIM may include a visual touch screen adapted to display a menu including icons, scroll bars, boxes, and buttons through which the operator may interface with the automated sample handling system <NUM> and wherein the menu comprises a number of function buttons programmed to display functional aspects of the automated sample handling system <NUM>.

In the instance described hereinabove wherein analyzer <NUM> is, for example, a clinical chemistry analyzer <NUM> and analyzer <NUM> is a coagulation analyzer, different centrifuge protocols may be established within centrifuge <NUM> in order to provide a properly centrifuged and pre-assay treated sample for testing by the chemistry analyzer <NUM> or by the coagulation analyzer <NUM>. As previously mentioned, sample containers <NUM> may be provided with identification indicia or information 20C readable by sensor <NUM> indicating the assay procedures to be accomplished upon the sample therein. Computer <NUM> is programmed to determine whether an assay is a clinical chemistry analysis or a coagulation analysis and which analyzers <NUM>, <NUM>, and <NUM> are adapted to perform such analyses.

<FIG> is schematic plan view of an apparatus which may be used at a sample quality station <NUM> of <FIG> and which may be adapted for automatically determining a presence of two or more interferents in a specimen 20A to be automatically processed by the automated sample handling system <NUM> of <FIG>. The presence of the interferents may be detected by the sample quality station <NUM> prior to being further tested by the automated sample handling system <NUM>. In this manner, if the specimen 20A includes an interferent, additional processing, discarding, or a redraw may take place. Additionally, other detection methods (not part of the invention) may take place on the analysis specimen 20A contained in the sample container <NUM>, as well as on the sample container <NUM> itself. For example, the apparatus may be used to determine certain physical dimensional characteristics of the specimen (e.g., the liquid-air interface LA, location of the interface SR between the red blood cell portion 20RBC and the serum or plasma portion 20SP, a height of the red blood cell portion HRB c, and a height of the serum plasma portion HS p, as shown in <FIG> and/or certain physical dimensional characteristics (e.g., height H or width W) of the sample container <NUM> (e.g., of the sample tube) or the location of the tube/cap interface TC.

Now referring to <FIG>, sample quality station <NUM> comprises a radiation source <NUM> (e.g., a collimated light source), for example. The radiation source <NUM> may be a laser diode, which directs a laser beam 52A of coherent light onto the sample container <NUM> (e.g., sample tube). The sample container <NUM> may be supported upon a rotating holder <NUM>, which may be a table or platform or other rotating apparatus adapted to support and hold the sample container <NUM> in a generally upright condition during rotation. Appropriate laser diodes for the radiation source <NUM> are well known in the art and include, inter alia, He, Ne, Gas, GAAS diodes. A radiation capture device <NUM>, such as a conventional digital camera, a charged coupled device (CCD), or a spectrophotometer, captures a beam of radiation transmitted through sample container <NUM>. The captured image of the transmitted beam of radiation is then analyzed to determine the presence of one or more interferents of the specimen 20A.

Sample quality station <NUM> further comprises a conventional light (radiation) source <NUM> of non-collimated visible light (e.g., white light) that also directs a beam of light 58A onto the sample container <NUM>. The light is captured by a radiation capture device <NUM> such as a conventional digital camera digitizing means, or an array of photodetectors, after transmission through sample container <NUM>. Reflectors and/or diffusers <NUM> may be provided to shape and obtain a uniform field of radiation. The radiation capture device <NUM> has a digitizing means, which is also preferably monochromatic with at least eight bit high accuracy analog to digital capability. A digitizing means meeting these requirements is the model DT <NUM> FrameGrabber, available from Data Translation. The digitized image is then analyzed with a computer (e.g., a controller, processor, or microprocessor) containing an image processing software program to determine the presence of two or more interferents in the specimen 20A.

The scattering of coherent light by a small, single particle is equivalent to transforming the morphology of the particle into the angular distribution of scattered light. Mathematically, this transform is known as the Fourier transform. This is a well-known effect and, for some classes of particles such as spheres, the transform is known exactly. When a collimated beam of coherent light encounters a specimen of biological cells or structures, part of the light may be absorbed, part may be scattered, and the rest may be transmitted. Transmitted and scattered light are measured in accordance with the invention to obtain certain information about the specimen 20A.

The scattering of coherent light by an ensemble of particles that are all within the coherence length of the light source is obtained by the superposition of the light scattered by individual particles. Thus, if the particles are uniform and dispersed randomly, the net scattering distribution is N times the scattering intensity of a single particle where N is the number of particles. This is also a well-known effect that has been used to both size and estimate a concentration of particles. These measurements may be made directly on the angular distribution of the scattered light or after performing a Fourier transform of the angular distribution. It has been discovered that particles such as fatty deposits of lipid present in the serum plasma portion 20SP have such an effect.

If the particles are not randomly dispersed, then the scattering from individual particles "interfere," creating a distribution of scattering intensity reflecting the organization of the individual particles. As a result, the tendency of particles to aggregate may be determined by measuring a fluctuation in intensity of the scattered light. If the particles are random, the "interferences" between the scattering from individual particles are also random, leading to a uniform and constant fluctuation at any angle. However, as particles begin to aggregate, the interference grows, leading to increased fluctuation that may be measured.

The specimen analysis process may start when a phlebotomist, physician, nurse, or health practitioner draws or collects a sample of blood from a patient and inserts the sample (specimen 20A) in a sample container <NUM>. That sample container <NUM> (e.g., sample tube) of blood is sent to a hospital laboratory, blood diagnostic laboratory, or other location to be tested for health conditions such as: diseases, cancers, fertility, drugs of abuse, and other disorders. The specific test for a health condition(s) depends on the patient and there are hundreds of health conditions that may be assessed using blood diagnostic instruments and automated analytical testing systems in these laboratories and locations. Blood diagnostic instruments, or clinical analyzers, may analyze the blood samples and provide the analytical results identifying the presence or an amount of a component therein. Automated systems, such as described in <FIG>, are used to transport the samples to and from blood diagnostic analyzer instruments <NUM>, <NUM>, <NUM>, as well as processing equipment such as centrifuge <NUM>.

Hospital or blood diagnostic laboratories can typically handle thousands of patient specimens each day and are increasingly relying on instruments and automation systems to process these samples accurately and efficiently. However, the instruments can produce errors that result from defects in the quality of the sample of blood. Sample quality defects may include discoloration, unwanted particle presence, clotting, and/or insufficient sample volume inside the sample tube.

Most of these defects can be observed after the centrifugation process. The centrifugation process in centrifuge <NUM> may separate the red blood cells (e.g., the red blood cell portion 20RBC) from the serum or plasma portion 20SP, where the red blood cells are packed at the bottom of the sample container <NUM>, and the serum or plasma portion 20SP (clear or relatively clear section) is at the top section of the sample container <NUM> (e.g., sample tube). The serum or plasma portion 20SP is where most of the defects such as discoloration, unwanted particle presence, and clotting may be observed.

In a non-automated lab, technicians visually and manually inspect the sample container <NUM> (e.g., sample tube) to determine whether a sample quality defect exists. In busy labs, that might not be done thoroughly or, due to the subjective nature of the inspection, the outcome may be highly variable. For example, there may be variations due to person-to-person inspection or even variations from one sample to the next sample when an individual person is inspecting.

In an automated lab, the samples are often automatically sent from the centrifuge to the analytical instruments (e.g., clinical analyzers), via an automation system, without the opportunity for inspection whatsoever. Many unwanted, problematic, and frustrating occurrences may be encountered when specimens 20A including these defects are provided to the one or more analytical instruments. For example, if the sample quality issue is discoloration or insufficient sample volume, the lab technician may have to remove the specimen 20A from the instrument and/or automation system and request a new sample of blood from that particular patient. This decreases efficiency in the lab and can be annoying to the patient because more blood has to be drawn from them, which may require additional visits to the phlebotomist.

In another example, if the sample quality issue is unwanted particle presence and/or clotting, the lab technician may have to perform additional preprocessing tasks for that sample before reinserting it into the system again for analysis. Additionally, the technician may have to stop the analytical instrument and/or automation system and clean certain equipment, such as probes and pipettes, on the system because of the particles that may have interfered with the assay (test).

Therefore, when any sample quality defect is encountered on an instrument, the assay results for that specimen may be erroneous and the efficiencies of the lab, such as test turnaround time or down time, may be negatively affected. The specimen 20A itself may also, in some instances, be wasted because the sample is analyzed before it has received the appropriate processing.

According to one sub-aspect, the method and apparatus (e.g., device) may detect a lipemic specimen of a centrifuged sample container (e.g., sample tube) containing a specimen (e.g., blood) using a collimated light source, preferably a laser diode, and a radiation capture device (e.g., a digital camera) for enabling electronic image analysis and detection.

Lipemia is a specific sample quality discoloration defect, which may be resolved with special processing before the specimen 20A is tested or analyzed on an analytical instrument or placed on an automation system. The definition of lipemia (also spelled lipaemia) is the abnormally high presence of lipids (fats) in the blood. Lipids exist as small particles not soluble in water. Typically, the serum or plasma portion 20SP (<FIG> is relatively clear. In a lipemic sample, however, the serum or plasma portion 20SP of centrifuged blood may appear to be white or milky in color due to the presence of the lipids. A common cause of lipemia is eating fatty foods. After the lab is aware the sample is lipemic, they may further process the specimen 20A to remove or reduce the lipids. For example, they may introduce a solvent or other material to reduce the amount of lipemia. Once this is complete, the specimen 20A can be properly analyzed by the clinical analyzer instrument (e.g., <NUM>, <NUM>, and <NUM>) and the lab will be relatively more confident of the test results.

This aspect seeks to detect lipemia at the first possible instance (e.g., at the next processing station) after centrifugation of the specimen 20A. By detecting lipemia at that point in the process, the specimen 20A will not be wasted, erroneous test results will be prevented, and the patient test result delay will be minimized. When this sample quality station <NUM> is provided on a automated sample handling system <NUM>, each specimen 20A will be screened for interfering levels of lipemia when it leaves the centrifuge <NUM> (See <FIG>. If an interfering level of lipemia is detected, the technician or user is alerted via a screen warning, warning bell, etc. The sample container <NUM> may then be routed to a place on the system <NUM> to wait for user corrective action or additional processing, such as to auxiliary processing station 30A. After the specimen 20A is corrected or additionally processed, it can be placed directly on an analytical instrument (e.g., <NUM>, <NUM>, and <NUM>) for analysis, or back onto the track <NUM> of the automated sample handling system <NUM>. The automated sample handling system <NUM> might be able to perform this corrective action on a sample without user interaction. For example, the routing of the lipemic specimen would remove the specimen via robotic transport 30B and require additional processing at station 30A as a prerequisite to analysis on the analytical instruments <NUM>, <NUM>, and/or <NUM>, for example.

According to the method <NUM>, and referring to <FIG>, <FIG>, a step of transmitting a beam of radiation 52A through the clinical analysis specimen 20A contained in the sample container <NUM> (e.g., sample tube) in block <NUM> may be by using a radiation source <NUM>, which may be a collimated light source, preferably generated by a laser diode, to project the beam 52A (e.g., a laser beam) onto a sample container <NUM> containing centrifuged blood having a red blood cell portion 20RBC and a serum or plasma portion 20SP. In the case where the radiation source <NUM> is a laser, the laser may be operated at any suitable wavelength and power. However, either <NUM> or <NUM> at <NUM> mWA are suitable. A radiation capture device <NUM>, such as a digital camera, CCD, or other suitable digitizing device, may capture an image (e.g., a digital image) of the sample container <NUM> of blood along with an image of the beam of radiation 52A as transmitted onto or through the sample container <NUM> in block <NUM>. A computer <NUM> containing a computer software program is used for electronic image analysis of the digital image captured by the image capture device <NUM>. The captured beam passing through the specimen 20A is analyzed for the presence of an interferent within the clinical analysis specimen 20A in block <NUM>. For example, the analysis may involve the detection of lipids as an interferent, for example. When the computer software successfully detects that a sample is lipemic, then that specimen 20A may not be immediately analyzed, and may be rerouted to another area on the instrument or automation system that is reserved for lipemic sample pre-processing activities (e.g., auxiliary processing 30C). The lab technician may then perform pre-processing activities and reinsert the specimen 20A on the analyzer instrument <NUM>, <NUM>, <NUM> or automated sample handling system <NUM>.

The sample container <NUM> may be located inside a non-reflective box or enclosure <NUM> for optimal image capture. Also located inside the enclosure <NUM> is the radiation source <NUM>, as well as another light source <NUM> and diffusers <NUM> that may serve to properly illuminate the sample container <NUM> for additional quality image capture results (e.g., for a determination of hemolysis). The radiation capture device <NUM> (e.g., digital camera) may be located either inside of the enclosure <NUM> or outside of the enclosure <NUM> and receive the image through the aforementioned view window of the sample container <NUM>. The sample container <NUM> may be positioned on a rotating holder <NUM>, which may be controlled by a motor (e.g., a stepper motor not shown) to rotate the sample container <NUM> until a visible region (a window where a label 20D is not applied) of interest is displayed so that the radiation capture device <NUM> may capture the beam of radiation (possibly diffused or deflected) to determine a presence of an interferent within the clinical analysis specimen 20A. The captured radiation is captured through the entire sample container <NUM> as in <FIG>. A spot size of the captured image of the beam is measured and correlated to a degree of lipemia present.

Specifically, lipids may interfere with spectrophotometric measurements of the analytical instruments mainly because they cause the light beam, used to measure absorbance in a sample within such instruments, to scatter. The scattered light is then not picked up by a radiation capture device of the spectrophotometer of the analytical instrument. Because the scattered light is not measured, it is assumed that the light was absorbed. Therefore, this will cause inaccurate analytical measurements.

In a lipemic sample, the property of light scattering that was described to cause problems for spectrophotometry during analysis is the very property that will be taken advantage of for measuring a lipid interferent. It was observed by the inventor that, when a beam 52A, 252A of radiation from a radiation source <NUM>, <NUM> (e.g., laser) passes through the serum or plasma portion 20SP of a normal specimen 20A, there is some light reflection from both the front and back surfaces of the sample container <NUM>, and some minor reflection from the specimen 20A itself, as is shown in <FIG>. On a sample with high concentration (i.e., which are lipemic), there is some light reflection from the front surface but, as the laser beam 52A, 252A enters the sample (specimen), there is immediate dispersion (scattering) and light reflecting from the specimen 20A, as shown in <FIG>. These same pictures with some simple image processing may further highlight the distinctive difference a lipemic sample has when exposed to a radiation source <NUM>, <NUM> (e.g., a laser). Further vision analysis and use of line generating optics to project a beam line (e.g., a laser line beam) onto the sample container <NUM> and specimen 20A may provide improved detection by allowing detection of the interfaces between air and serum or plasma portion 20SP in the container <NUM> (not part of the invention), as well as an interface SR between the serum or plasma portion 20SP and the red blood cell portion 20RBC (not part of the invention). Additionally, an interface TC between the sample tube 20T and cap 20B of the container <NUM> may be determined (not part of the invention).

The method <NUM> includes providing a radiation source <NUM> (<NUM>) that transmits a beam of radiation onto the sample container <NUM> containing the specimen 20A in block <NUM>, and then in block <NUM>, a radiation capture device <NUM> (<NUM>) may capture an image of the sample container <NUM> with beam 52A (252A) including reflections and/or dispersions superimposed thereon in block <NUM>. The image may contain a laser beam spot <NUM> , <NUM> as shown in <FIG> illustrates a normal spot size <NUM> and <FIG> illustrates an image of a lipemic sample including a relatively enlarged spot size <NUM>. For example, the spot <NUM> may be enlarged having a width dimension W2, as compared to a normal sample having a spot <NUM> with a smaller width dimension W1. Processing software may measure a size of the spot <NUM> , <NUM> and correlate that spot size to a degree of dispersion caused by the specimen 20A as the beam 52A (252A) disperses within the specimen 20A. This, in turn, suggests a lipid concentration. Generally, the only substances that cause such dispersion in a specimen 20A are lipids but, regardless of the substance, if the light going through the specimen 20A is appreciably dispersed, an analytical measurement error is likely, and the lab should be appropriately informed, such as by generating a screen warning, sounding an alarm, unloading the lipemic specimen 20A, or stopping the automated system <NUM>. As discussed above, the relative amount of dispersion, which may be dictated by the spot size (e.g., a spot width) may be correlated to a lipemic index. If the lipemic index exceeds a preset threshold, the specimen 20A may be determined to be lipemic. In the case where line generating optics are used, the captured image (e.g., reflected image) may also be used to determine the presence of one or more interferents by examining the portions of the image. Additionally, since the presence of lipemia is generally indicated by a white color, according to the invention the serum or plasma portion 20SP is further analyzed using the radiation source <NUM> and capture device <NUM> to generate an image thereof. RGB values (a red, green, blue system of color analysis) obtained from the captured image may be analyzed by comparing them to stored threshold values (indicative of a certain hue of white) to indicate the presence of lipemia.

At least five things make the lipemia measurement technique relatively cost effective. One is that the pre-screen consumes none of the specimen 20A; it does not require an open sample container <NUM> or need to come in contact with the specimen 20A in any way. Two is that it is fast; the image acquisition and analysis may be significantly faster than chemistry analysis techniques. Three is that there is no variable cost; with no consumables, the cost per sample analyzed is substantially zero. Four is that the pre-screen may be performed early in the pre-analytical processing phase of specimen preparation; any necessary corrective action may be undertaken on the specimen 20A before clinical analysis is attempted. Five is that the pre-screen may measure directly, without any chemical analysis, the phenomenon dispersion that may cause interference in the spectrophotometer.

Additionally, but not as part of the invention, the capture image may be used to identify such items as: <NUM>) the vertical interface location TC between the sample tube 20T and sample cap 20B, which may establish a relative height (H) of the sample container <NUM> being used; <NUM>) the vertical interface location SR between the serum or plasma portion 20SP and the red blood cell portion 20RBC; and <NUM>) the vertical interface location LA between the serum or plasma portion 20SP and the air 20E in the sample container <NUM>.

According to another sub-aspect, the method and apparatus (e.g., device) may be used to detect a hemolyzed sample (specimen) contained in a sample container <NUM> of centrifuged blood. The method <NUM> as shown in <FIG> (not part of the invention) utilizes a radiation source for projecting light radiation (e.g., a white light) onto the sample container <NUM>, a radiation capture device (e.g., a digital camera) for electronic image capture, and then analysis of the captured image to detect hemolysis.

Hemolysis is a sample quality discoloration issue, and it cannot be resolved with special processing. Hemolysis (also spelled haemolysis) may occur when the red blood cells rupture and the hemoglobin inside is released into the serum/plasma section 20SP (<FIG> of the centrifuged blood specimen 20A, thus giving the serum or plasma section 20SP a more reddish color or appearance. Along with a more reddish color, potassium may be released into the serum or plasma portion 20SP, which may give erroneous results when tested on an analytical instrument. Incorrect blood collection, handling, storage, and/or processing may cause hemolysis.

In the case of a clinical analysis specimen 20A suffering from hemolysis, the usual procedure is to redraw another specimen 20A from the patient to ensure that a good quality specimen 20A is presented to the analytical instrument(s). Once the new specimen 20A is processed, it may be successfully analyzed without the interfering hemoglobin.

In another aspect (not part of the invention), one can detect the presence of hemolysis in the specimen 20A, thereby saving the analytical instrument(s) from performing analytical testing on a specimen 20A whose results may be suspect. When the specimen 20A is imaged and analyzed for lipemia, the serum or plasma portion 20SP is visible through the side (window) of the sample container <NUM> (e.g., test tube). At this time, at the quality control station <NUM>, there is an opportunity to record and analyze the color of the serum or plasma portion 20SP in order to make a basic assessment for hemolysis. This assessment may be solely performed or may be performed in conjunction with the lipemic analysis at the quality control station <NUM>.

In the assessment for hemolysis, the radiation source <NUM> (e.g., collimated light source or laser) would be turned off. As described above, the "window," or region of interest, where there is no label 20D may have already been found via rotation of the sample container <NUM> during the earlier search, such as when analyzing for potential lipemia. If not, the procedure above may be employed wherein in block <NUM> the sample container <NUM> is rotated to orient the window relative to a radiation source and appropriately position the sample container <NUM> for hemolysis assessment. Additionally, with image capture and processing for hemolysis analysis, the area under consideration is reduced to just the serum or plasma portion 20SP in the "window.

Unlike a lipemic specimen, when the computer software successfully detects that a specimen 20A is hemolyzed, then that specimen 20A may continue to be tested and analyzed on the clinical analytical instrument(s) without delay. After completion of the analytical testing, the specimen 20A may be rerouted to an area on the instrument or the automated sample handling system <NUM> that is reserved for evaluating hemolyzed samples. For any successful detection of a hemolyzed sample, the computer may provide an alert that will be displayed on a display (e.g., computer screen) at the quality control station <NUM>, on the analytical instrument, or automated sample handling system <NUM> to alert personnel (e.g., technician) for further evaluation and/or decision making.

To improve an ability to convey the assessment of a hemolyzed sample to laboratory personnel, an image of the sample container <NUM> (e.g., test tube) including the specimen 20A determined to be hemolyzed may be displayed. This may be displayed along with collaborative information such as, but not limited to, reference images of various hemolyzed specimens, color spectra for comparison, sample's assessed position in color spectra, and/or text description of issue and/or suggested laboratory action to take.

In addition, if a hemolyzed specimen 20A were detected on a sample quality station <NUM> of an automated sample handling system <NUM>, the specimen 20A may be sent on to an analytical instrument (e.g., a specialized clinical analyzer) where a precise level of hemolysis can be measured and characterized. Analytical instruments are much better at determining levels of hemolysis and often have rules that determine the exact concentrations of hemoglobin that affect assay results for the various assays ordered for the specimen. As a result, some test results can be reported before the specimen redraw and retesting occurs. However, it should be apparent that, with the early detection of hemolysis, the laboratory technician can decide on the urgency of a redraw with the confidence that the automated sample handling system <NUM> may be able to report the results, even though the specimen 20A may contain some level of hemolysis. Additionally, or optionally, an alert could also be used to identify which ordered assay results are likely to have an adverse effect by the extent of hemolysis that has been detected. With this enhanced presentation, the task of making the correct clinical decision may be made significantly easier and less prone to error.

The action to take when a sample is hemolyzed is based on rules defined by the laboratory to align with their specific procedures. The hemolysis threshold that triggers the rules may also be established by the laboratory, and may vary from test to test, which may also be subsequently undergone. The lab would use the quality control specimen (e.g., a reference sample 20R - see <FIG>) to establish a threshold hemolysis level that triggers the alert. For example, sample 20R shown in <FIG> illustrates a reference sample having a serum or plasma portion 20SP with an elevated level of hemolysis. Sample <NUM> in <FIG> illustrates a sample with an elevated level of hemolysis above the threshold. An example of various levels of hemolysis may be seen in an article in <NPL>.

Now referring to <FIG> and <FIG>, in order to determine an amount of hemolysis present in the specimen 20R (<FIG>) contained in the sample container <NUM>, a radiation source <NUM> projects a light beam 58A onto the sample container <NUM> in block <NUM>, and an image is captured by the radiation capture device <NUM> in block <NUM>. This image is analyzed in block <NUM> to determine a color of the serum or plasma portion 20SP. A red, green, blue (RGB) system of color analysis may be employed. Accordingly, the light capture device <NUM> may be any suitable camera or digitizing array capable of discerning RGB hues. The respective red (R) hue may be measured on a scale from <NUM> to a maximum number (e.g., <NUM> -<NUM>). Any specimen 20A, which may include a red hue above a threshold value as established by calibrating with the reference sample 20R, may be determined to be a hemolyzed sample. Optionally, more than one color may be measured and thresholds may be set based upon more than one detected color. Image analysis in block <NUM> may include measuring the color of the serum or plasma portion 20SP in an area located vertically between the liquid-air interface LA and the interface SR, and roughly centered on the window. Based upon the detected hue or hues, a hemolytic index may be determined and reported, or otherwise conveyed from the quality control station <NUM>. The hemolytic index may be roughly determined based upon correlated ranges of the measured hue values as shown in Table <NUM> below, for example.

The hemolysis measurement technique shares most of the advantages indicated above for the lipemic detection. One advantage is that the hemolysis pre-screen consumes no specimen 20A; it does not require an open sample container <NUM> or need to come in contact with the specimen 20A in any way. Two is that it is relatively fast; the image acquisition and analysis may be significantly faster than chemistry analysis techniques. Three is that there may be substantially no variable cost; with no consumables, the cost per specimen analyzed is substantially zero. Four is that the hemolysis pre-screen may be performed early in the pre-analytical processing phase of specimen preparation; hospital personnel can be alerted as early as possible that there is a condition of the specimen 20A that may require their attention.

According to another sub-aspect, the method and apparatus (e.g., device) may be used to detect icterus in a specimen 20A contained in a sample container <NUM> of centrifuged blood. An icterus interferent may arise, for example, from an excess of bilirubin, the result of decaying red blood cells being converted in the spleen into bilirubin. Levels of bilirubin above <NUM>-<NUM>/dl are generally visibly yellowish/brownish in color and may, in particular, adversely affect enzyme-based immunoassays. Such a condition is also termed bilirubinaemia.

The icterus detection method 500A (see <FIG>) is similar to that for detecting hemolysis. The method 500A may first rotate the sample container to orient the window in block 502A. Next, the method 500A utilizes a radiation source (e.g., radiation source <NUM>) for transmitting a beam of light radiation 58A (e.g., white light) onto the sample container <NUM> containing a clinical analysis specimen 20A in block 504A. The apparatus may be as shown in <FIG>, for example. A radiation capture device (e.g., <NUM> such as a digital camera) adapted for digital electronic image capture captures an image from the clinical analysis specimen 20A of the beam 52A as passing through the sample container <NUM> and serum or plasma portion 20SP of the specimen 20A in block 506A. A computer <NUM> may then perform an analysis of the captured image for the presence of icterus. According to the method, the same digital image that was taken for the hemolysis detection may optionally be used for icterus detection. In this case, the image may be analyzed for the presence of a yellow and/or brown color. Again this may be accomplished via measuring with the radiation capture device <NUM> (e.g., a digital camera having RGB capability or an RGB sensor) a degree of yellow and/or brown present in the serum or plasma portion 20SP of the specimen 20A. Optionally, a sensor using the CMYK system may be employed. Range values for each of yellow and/or brown may be experimentally determined and set and may be used to provide an icteric index. For example, a range from <NUM> to <NUM> may be employed. Other suitable icteric index values may be used. A central portion of the serum or plasma portion 20SP for analysis may be located via the image analysis technique described herein for determining the location of interfaces LA and SR.

There are several alternative embodiments of this invention. In one embodiment shown in <FIG> and described above with reference to <FIG>, line generating optics 252B are coupled to a radiation source <NUM> (e.g., a collimated source such as a laser) to project a laser beam 252A in the form of a "line" onto the sample container <NUM> and specimen 20A. In another embodiment as shown in <FIG>, the radiation source <NUM> may be moved vertically along a vertical axis aligned with the Z axis and simply sweep a spot of the laser beam longitudinally along the sample container <NUM> and specimen 20A between limits A, B. In this embodiment, the width, vertical location, and horizontal location of the spot at various vertical positions may be measured by a rotationally offset capture device <NUM> as described above. This data may be correlated to be able to locate the tube-cap interface TC, liquid-air interface LA, the SR interface, the SS interface, the SSR interface (all not part of the invention), as well as the presence of lipemia.

Claim 1:
An apparatus adapted to determine the presence of at least two interferents in a clinical analysis specimen contained within a sample container (<NUM>), comprising:
a radiation source (<NUM>) adapted to transmit a beam (52A) of radiation through the clinical analysis specimen contained within the sample container (<NUM>);
a first radiation capture device (<NUM>) adapted to capture said beam of radiation as transmitted through the sample container (<NUM>); and
a light source (<NUM>) adapted to transmit a conventional light beam (58A) of non-collimated visible light through the clinical analysis specimen contained within the sample container (<NUM>);
a light capture device adapted to capture the light beam of non-collimated visible light transmitted through the sample container (<NUM>), said light capture device comprising a second radiation capture device (<NUM>), which has digitizing means; and
a computer (<NUM>) adapted to analyze the captured beam of radiation and the captured digitized beam of visible light to determine
from the analysis of each of said beams (52A, 58A) a presence of one or more interferents within the clinical analysis specimen.