Automated immunoassay cassette, apparatus and method
An immunoassay cassette, apparatus, and method for detecting an analyte in a liquid body-fluid sample are disclosed. The cassette has a body and a support mounted on the body, for movement toward and away from a sample transfer position. Sample supplied to a sample well in the cassette body is taken up by a reagent reservoir containing a first reagent composition effective to form a modified sample. The support provides a reagent strip having a transfer zone that is brought into contact with the reservoir, when the support is in the transfer position, a detection zone located downstream of the transfer zone, and a second reagent composition effective to react with the modified sample to form a detectable analyte-dependent product. By controlling movement of the support, volume and rate of sample flow from the reservoir to the strip can be controlled to optimize and/or standardize sample-transfer conditions in the assay.
1. An immunoassay cassette for use in detecting an analyte in a liquid body fluid sample, comprising:
(a) a cassette body having a sample well for receiving said sample;
(b) carried on said body, (i) a reservoir containing a first reagent composition effective to react with one or more sample components to form a modified sample, when sample migrates from the sample well into the reservoir;
(c) a support mounted on said body, for movement toward and away from a transfer position; and
(d) carried on said support, a reagent strip containing a second reagent composition effective to react with the modified sample formed in said reservoir to form a detectable analyte-dependent product, said strip having a transfer zone which is brought into contact with said reagent reservoir, when the support is moved to its sample-transfer position, and a detection zone located downstream of the transfer zone,
wherein controlled movement of said support toward and away from its transfer position is effective to meter the volume and rate of sample flow from the reservoir to the reagent strip.
2. The cassette of claim 1 , for detecting a multivalent analyte in a liquid body-fluid sample, wherein the first reagent composition in the reagent reservoir includes a non-immobilized conjugate of an anti-analyte antibody and a detectable reporter group, the reaction to form a modified sample includes binding of the conjugate to sample analyte, to form an analyte-conjugate complex, the reagent composition in the reagent strip includes an anti-analyte antibody immobilized at a detection region in the reagent strip, and the reaction to form a detectable analyte-dependent product includes binding of complex to the immobilized antibody, to localize the detectable reporter in the complex at the detection zone.
3. The cassette of claim 2 , wherein said non-immobilized conjugate is a conjugate of an anti-analyte antibody and a detectable reporter selected from the group consisting of metal particles, particles labeled with colored or fluorescent moieties, polymers labeled with colored of fluorescent moieties, particles, and colored or fluorescent molecules.
4. The cassette of claim 3 , for use in detecting C-reactive protein analyte in a blood sample, wherein the anti-analyte antibody in the non-immobilized conjugate in the reagent reservoir, and the immobilized anti-analyte antibody in the reagent strip are antibodies specific against a common epitope in C-reactive protein.
5. The cassette of claim 1 , wherein said support includes a window through which a detectable reaction at the detection zone in the reagent strip be viewed.
6. The cassette of claim 4 , wherein the detection zone in the reagent strip is covered by a reflective film at the strip's surface facing away from said window, such that flow of sample liquid through the detection zone produces a first change in reflectance measurable through the window, and the presence of analyte-dependent reaction at the detection zone produces a second change in reflectance measurable through the window.
7. The cassette of claim 1 , which further includes an absorbent reservoir carried on said support, downstream of said detection zone, and in fluid-flow communication with said reagent strip, for receiving sample liquid transferred onto the reagent strip.
8. An apparatus for use in detecting an analyte in a liquid body-fluid sample, comprising:
A) a cassette having
(a) a cassette body having a sample well for receiving said sample,
(b) carried on said cassette body, (i) a reagent reservoir containing a first reagent composition effective to react with one or more sample components to form a modified sample, as sample migrates from the sample well into the reservoir,
(c) a support mounted with respect to said cassette body, for movement toward and away from a transfer position,
(d) carried on said support, a reagent strip containing a second reagent composition effective to react with the modified sample to form a detectable analyte-dependent product, said support containing a sample-transfer zone which is brought into contact with said reservoir, when the support is moved to its transfer position, and a detection zone downstream of the transfer zone; and
(B) a cassette handling instrument having
(a) a cassette holder into which the cassette is removably placed, during a sample assay,
(b) an actuator operable to move the support in the cassette toward and away from its sample-transfer position,
(c) a detector operable to detect an analyte-specific reaction at the detection zone in the reagent strip, and
(d) a control unit operably connected to the actuator, for controlling the volume and rate of sample transfer from said reagent reservoir to the reagent strip during an assay procedure.
9. The apparatus of claim 8 , for detecting a multivalent analyte in a liquid body-fluid sample, wherein the first reagent composition in the reagent reservoir includes a non-immobilized conjugate of an anti-analyte antibody and a detectable reporter group, the reaction to form a modified sample includes binding of the conjugate to sample analyte, to form an analyte-conjugate complex, the reagent composition in the reagent strip includes an anti-analyte antibody immobilized at a detection region in the reagent strip, and the reaction to form a detectable analyte-dependent product includes binding of complex to the immobilized antibody, to localize the detectable reporter in the complex at the detection zone.
10. The apparatus of claim 9 , wherein said non-immobilized conjugate is a conjugate of an anti-analyte antibody and a detectable reporter selected from the group consisting of metal particles, particles labeled with colored or fluorescent moieties, polymers labeled with colored of fluorescent moieties, particles, and colored or fluorescent molecules.
11. The apparatus of claim 10 , for use in detecting C-reactive protein analyte in a blood sample, wherein the anti-analyte antibody in the non-immobilized conjugate in the reagent reservoir, and the immobilized anti-analyte antibody in the reagent strip are antibodies specific against a common epitope in C-reactive protein.
12. The apparatus of claim 8 , wherein said support includes a window through which a detectable reaction at the detection zone in the reagent strip be detected by said detector.
13. The apparatus of claim 12 , wherein the detection zone in the reagent strip is covered by a reflective film at the strip's surface facing away from said window, such that flow of sample liquid through the detection zone produces a first change in reflectance measurable through the window, and the presence of analyte-dependent reaction at the detection zone produces a second change in reflectance measurable through the window.
14. The apparatus of claim 13 , wherein said detector is operable to detect liquid flow through the detection zone, by a first change in measured optical reflectance, and is operable to measure a subsequent analyte-dependent reaction at the detection zone, by a second change in measured optical reflectance.
15. The apparatus of claim 8 , wherein said control unit is operable to control the volume and rate of sample transfer from the reagent reservoir to the reagent strip by controlling one or more of (i) the period of sample incubation before sample is first transferred from the reservoir to the reagent strip, (ii) the cycle frequency with which the actuator moves the support toward and away from its transfer position, (iii) the time of contact that the support is held in its transfer position, during each cycle, and (iv) the total number of transfer cycles.
16. The apparatus of claim 15 , wherein the control unit is operable to control the rate of sample transfer from said reservoir to the reagent strip by controlling (i) the cycle frequency with which the actuator moves the support toward and away from its transfer position and (ii) the time of contact that the support is held in its transfer position, during each cycle.
17. A method of conducting an assay for a body-fluid analyte comprising:
introducing a body fluid containing the analyte into an absorbent reservoir containing a first reagent composition effective to react with one or more sample components to form a modified sample;
repeatedly contacting the reservoir, with such reservoir containing an absorbed body-fluid sample, with a reagent strip containing a second reagent composition effective to react with the modified sample formed in said reservoir to produce a detectable analyte-dependent product; and
controlling the frequency and duration of said repeated contacting, thereby to control the volume and rate of transfer of sample fluid from the reservoir to the pad.
18. The method of claim 17 , wherein said reagent strip is an elongate reagent strip having a sample-transfer zone at which the reservoir makes contact with the strip, and a detection zone located downstream of the transfer zone.
19. The method of claim 18 , for detecting a multivalent analyte in a liquid body-fluid sample, wherein the first reagent composition in the absorbent reservoir includes a non-immobilized conjugate of an anti-analyte antibody and a detectable reporter group, the reaction to form a modified sample includes binding of the conjugate to sample analyte, to form an analyte-conjugate complex, the reagent composition in the reagent strip includes an anti-analyte antibody immobilized at a detection region in the reagent strip, and the reaction to form a detectable analyte-dependent product includes binding of complex to the immobilized antibody, to localize the detectable reporter in the complex at the detection zone.
20. The method of claim 19 , wherein said non-immobilized conjugate is a conjugate of an anti-analyte antibody and a detectable reporter selected from the group consisting of metal particles, particles labeled with colored or fluorescent moieties, polymers labeled with colored of fluorescent moieties, particles, and colored or fluorescent molecules.
21. The method of claim 20 , for use in detecting C-reactive protein analyte in a blood sample, wherein the anti-analyte antibody in the nonimmobilized conjugate in the reagent reservoir, and the immobilized anti-analyte antibody in the reagent strip are antibodies specific against a common epitope in C-reactive protein.