Use of particulate labels in bioanalyte detection methods

• US 7,413,868 B2
• Filed: 11/03/2004
• Issued: 08/19/2008
• Est. Priority Date: 11/05/2003
• Status: Expired due to Fees

• Current Assignee: Trellis Bioscience LLC
• Original Assignee: Trellis Bioscience LLC
• Inventors: Cromer, Remy, Kauvar, Lawrence M., Collarini, Ellen J., Harriman, William D.
• Primary Examiner(s): GABEL, GAILENE
• Application Number:
  US10/981,130
• Publication Number:
  US 20050106641A1
• Time in Patent Office: 1,385 Days
• Field of Search: 435/7.1, 435 721- 724, 435/7.92, 435/7.94, 435/372.2, 435/372.3, 435/287.9, 435/287.2, 435/968, 435/973, 436/518, 436/521, 436/522, 436/77, 436524-530, 436/533, 436/546, 436/10, 436/17, 436/63, 436/64, 436/172, 436/46, 436/56
• US Class Current: 435/7.2
• CPC Class Codes: G01N 15/0625 Optical scan of the deposits
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 2030/8813 biological materials
• CPC Class Codes: G01N 2030/8827 involving nucleic acids
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6458 Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• CPC Class Codes: G01N 27/44726 using specific dyes, markers or binding molecules
• CPC Class Codes: G01N 30/74 Optical detectors measurement of intensity, velocity, spectral content, polarisation, or phase of infrared, visible or ultraviolet light G01J
• CPC Class Codes: G01N 30/88 Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
• CPC Class Codes: G01N 33/505 involving T-cells
• CPC Class Codes: G01N 33/5052 involving B-cells
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form
• CPC Class Codes: G01N 33/585 with a particulate label, e.g. coloured latex
• CPC Class Codes: Y10T 436/112499 with sample on test slide
• CPC Class Codes: Y10T 436/13 Tracers or tags
• CPC Class Codes: Y10T 436/25125 Digestion or removing interfering materials
• CPC Class Codes: Y10T 436/25375 Liberation or purification of sample or separation of material from a sample [e.g., filtering, centrifuging, etc.]

Abstract

New applications for the use of distinguishable particulate labels available in a variety of hues and sized in the submicron range are described. These applications include profiling of cellular components, obtaining secretion patterns, identifying a multiplicity of components in chromatographic or electrophoretic techniques and identification of desired immunoglobulin secreting cells.

Claims

1. A method to determine the presence, absence or amount of each of a multiplicity of components released from a single cell and/or the nature of at least one released component from the single cell, which method comprises permeabilizing or lysing the single cell or allowing the single cell to secrete cellular components,

disposing the released components, on a planar sample surface, contacting the released components disposed on said surface with a multiplicity of particulate labels having diameters of 10-500 nm with different hues, wherein each differently hued particle is conjugated to specific binding partner for a cellular component or portion thereof;

and

wherein particulate labels bound to components are retained but particulate labels not bound to components are not retained;

removing particulate labels not bound to components; and

observing the number and nature of individual particulate labels that are bound to the released components on the sample surface by means of microscopic observation,

whereby the presence, absence or amount of each of a multiplicity of released components and/or the nature of at least one released component from said single cell are determined.

2. The method of claim 1, wherein the amount of each different released component is assembled to obtain a profile of said cell with respect to said released components, wherein said profile is represented by a location in n dimensional space, wherein n is the number of components for which specific binding partner-containing particulate labels have been supplied.

3. The method of claim 2, wherein the cell is a T-cell and the released components comprise at least three different cytokines.

4. The method of claim 3, which is performed on at least 50 T-cells representative of the T-cell population of a subject for whom a disease state is to be assessed.

5. The method of claim 2, which further includes exposing said cell to a drug or other external stimulus prior to said contacting.

6. The method of claim 5, wherein said cell is obtained from a tissue sample.

7. The method of claim 5, wherein the number and nature of particulate labels observed under conditions where said cell has not been exposed to said drug or stimulus is compared to the number and nature of particulate labels observed when the cell has been exposed to said drug or external stimulus, thereby determining the effect of said drug or external stimulus on said cell.

8. The method of claim 1, wherein the released components are products of reporter genes.

9. The method of claim 1, wherein the cell is a T-cell and the released components comprise at least three different cytokines.

10. The method of claim 9, which is performed on at least 50 T-cells representative of the T-cell population of a subject for whom a disease state is to be assessed.

11. The method of claim 1, wherein the cell is a primary B-cell, an immortalized B-cell or a hybridoma and the component the nature of which is to be determined is an antibody specific for a specific antigen or epitope.

12. The method of claim 1, which further includes exposing said cell to a drug or other external stimulus prior to said contacting.

13. The method of claim 12, wherein said cell is obtained from a tissue sample.

14. The method of claim 12, wherein the number and nature of particulate labels observed under conditions where said cell has not been exposed to said drug or stimulus is compared to the number and nature of particulate labels observed when the cell has been exposed to said drug or external stimulus, thereby determining the effect of said drug or external stimulus on said cell.

15. The method of claim 1, wherein said cell is lysed or permeabilized.

16. The method of claim 1, wherein said released components are secreted components.

17. The method of claim 1, wherein said single cell is supported on a permeable membrane support which is overlaid on said sample surface on which said released components are deposited.

18. The method of claim 17 which is performed on an array of individual cells and wherein the location of the sample surface is correlated with the position of each cell on the membrane.