Methods for performing dry chemistry, lateral flow-reconstituted chromatographic enzyme-driven assays
View Patent ↗Methods for performing dry chemistry, lateral flow-reconstituted chromatographic enzyme-driven assays are described. A combination of components necessary to elicit a specific enzyme reaction are predeposited as substrate in dry form together with ingredients necessary to produce a desired color upon occurrence of the desired reaction. The strip is equipped with a sample pad placed ahead of the substrate deposit in the flowstream, to which liquid sample is applied. The sample flows from the sample pad into the substrate zone where it immediately reconstitutes the dried ingredients while also intimately mixing with them and reacting with them at the fluid front. The fluid front moves rapidly into the final “read zone” wherein the color developed is read against predetermined color standards for the desired reaction. The assay in the format of the invention is faster and easier to perform than analogous wet chemistry assays.
1. A method of assaying for an analyte in a blood sample, the method comprising:
(a) applying the sample to a chromatographic strip, the chromatographic strip comprising at least a first pad and a second pad, wherein said first pad is a sample receiving pad which has been treated with a lysing agent to lyse the sample as it flows therethrough, releasing hemoglobin from the sample and the second pad comprises a mixture of dry components; and
(b) without prior removal of the hemoglobin from the sample, allowing said sample to flow through the first pad and into the second pad, thereby mobilizing the mixture of dry components,
whereby the dry components are reconstituted by contact with the blood sample and a color indicative of the presence or concentration of the analyte is formed at a forward flow front of the chromatographic strip adjacent to a region colored by the hemoglobin from the sample.
2. The method of claim 1 , wherein the first pad is a sample receiving pad, wherein a region of the sample receiving pad has been treated to concentrate the sample as it flows therethrough by removing a portion of its liquid content.
3. The method of claim 1 , wherein the first pad is a sample receiving pad and the second pad is a substrate pad, wherein an intermediate pad is interposed between the sample receiving pad and the substrate pad.
4. The method of claim 1 , wherein the sample is obtained from a patient.
5. The method of claim 1 , wherein the color indicative of the presence or concentration the analyte can be evaluated within 5-20 minutes of applying the sample to the chromatographic strip.
6. The method of claim 1 , wherein the analyte is selected from the group consisting of glucose, galactose cholesterol, HDL-cholesterol, triglycerides, urea nitrogen, creatinine, glucose-6-phosphate dehydrogenase (G6PD), alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase, creatinine kinase, beta-lactamase, and peroxidase.
7. The method of claim 1 , wherein the analyte is an enzyme.
8. The method of claim 7 , wherein the enzyme selected from the group consisting of glucose-6-phosphate dehydrogenase (G6PD), alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase, creatinine kinase, beta-lactamase, and peroxidase.
9. The method of claim 7 , wherein the enzyme is G6PD.
10. The method of claim 1 , wherein the mixture of dry components in the second pad includes a substrate for an enzyme.
11. The method of claim 1 , wherein the region colored by hemoglobin in the sample is a region from which dried components have been removed from the second pad by the forward flow of the blood sample.