Colorable microspheres for DNA and protein microarray

A microarray and microsphere technology, applied in the field of biological microarrays, can solve the problems of fluorescence signal interference, fluorescence signal intensity suppression, limiting the diversity of color barcode coding of microspheres, etc. The effect of dynamic range

Inactive Publication Date: 2006-08-30
CARESTREAM HEALTH INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method has two problems: (1) the colorant itself emits fluorescence, which interferes with the fluorescent signal generated by the biological interaction; (2) when the barcode encodes the adsorption wavelength of the dye and the biological interaction When the fluorescence emission of the two is in a complementary relationship, the fluorescence signal intensity will be significantly suppressed
Problem 1 severely limits the diversity of microsphere color barcode coding, and Problem 2 significantly reduces the dynamic range and lower limit of detection of the microarray system

Method used

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  • Colorable microspheres for DNA and protein microarray
  • Colorable microspheres for DNA and protein microarray
  • Colorable microspheres for DNA and protein microarray

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0113] This example illustrates two methods for loading a photocoupler as a latent colorant into polystyrene microspheres.

[0114] Loading method 1: For the general preparation process, a single coupler, or more than one coupler in a fixed ratio, and different ratios of couplers, coupler solvents, and auxiliary coupler solvents are used to prepare microsphere samples. The cyan coupling agent CYAN1 was loaded by ultrasonic treatment, specifically as follows: 0.08 g of CYAN1 was stirred and dissolved in 0.8 g of cyclohexanone and 0.08 g of tricresyl phosphate (tricresolphosphate). This oil phase was then added to an aqueous phase of 0.48 g FAC-0064 (surfactant) and 6.52 g water with stirring at room temperature. The sample was sonicated for 1 minute to give a milky white dispersion, which was then stirred. An equal amount of 8.0 g of 4% 10 micron polystyrene microspheres was added to the sonicated sample. After mixing, the samples were poured into diafiltration bags and washe...

Embodiment 2

[0119] These examples illustrate the loading of photographic couplers as latent colorants into polystyrene microspheres using in situ polymerization.

[0120]

[0121] Table 1

[0122] Monomer-Coupler 1 Monomer-Coupler 2 Monomer-Coupler 3

[0123] (cyan) (yellow) (magenta)

[0124] beads#

1 (cyan)

2 (yellow)

3 (magenta)

Monomer-Coupler 1 (g)

0.85

-

-

monomer-coupling

Agent 2 (g)

-

0.85

-

monomer-coupling

Agent 3 (g)

-

-

0.85

Styrene (ml)

36.6

36.6

36.6

AIBN(g)

0.38

0.38

0.38

Ethanol (ml)

87.5

87.5

87.5

Methyl solution

Fiber

(ml)

125.0

125.0

125.0

Polyacrylic acid

(g)

3.75

3.75

3.75

Average

Grain diameter

(μm)

4.26

4.92

7.54

...

Embodiment 3

[0128] This example demonstrates the attachment of presynthesized single-stranded oligonucleotide probes to the surface of microspheres incorporating coupling agents.

[0129] Wash 100 μl of microspheres (4% w / v) incorporating coupling agent three times in acetate buffer (0.01M, pH 5.0), and mix with 100 μl of 20 mM 2-(4-dimethylcarbamoyl -pyrido)-ethane-1-sulfonate in combination with 10% polyethyleneimine. The mixture was stirred at room temperature for 1 hour and washed three times with sodium borate buffer (0.05M, pH 8.3). Resuspend the beads in sodium borate buffer.

[0130] Dissolve the 5'-amino-C6 modified oligonucleotide DNA probe in 100 μl of sodium borate buffer to a final concentration of 40 nmol. 20 µl of cyanuric chloride in acetonitrile was added to the DNA probe solution, and the total volume was brought up to 250 µl with sodium borate buffer solution. The resulting solution was stirred at room temperature for 1 hour, and then dialyzed against 1 liter of bori...

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Abstract

A microarray comprising: a support, on which is disposed a layer of microspheres bearing biological probes; wherein said microspheres comprise at least one material with a latent color that can be developed and used to identify said microsphere. A method of identifying biological analytes using the microarray is also disclosed.

Description

field of invention [0001] The present invention relates generally to biological microarray technology. In particular, the present invention relates to arrays of microspheres immobilized on substrates and methods of exposing the surface of the microspheres to analytes contained in a test sample. The microspheres contain a latent colorant that recognizes the microspheres when a color develops. The microspheres also carry capture agents (also called probes) on the surface. Background of the invention [0002] Various methods have been used in the prior art to prepare microarrays. For example, US Patent Nos. 5,143,854, 5,412,087, and 5,489,678 demonstrate that photolithography can be used to prepare peptide and DNA microarrays. This patent teaches the use of photolabile protecting groups to continuously cycle through photolithography to remove the protection of a defined point on a 1cm×1cm chip, and then fill the entire surface with active amino acids or DNA bases to prepare ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J19/00G01N33/58G01N33/543
CPCB01J2219/0072B01J2219/00545G01N33/54386B01J2219/00659B01J2219/00648G01N33/583B01J2219/00466B01J2219/00725B01J2219/00722B01J19/0046B01J2219/00731G01N33/54313B01J2219/00576
Inventor T·A·乔J·W·莱昂K·M·施勒德
Owner CARESTREAM HEALTH INC
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