Microarrays utilizing hydrogels

• US 7,595,157 B2
• Filed: 08/19/2004
• Issued: 09/29/2009
• Est. Priority Date: 08/19/2004
• Status: Active Grant

• Current Assignee: Biocept, Inc.
• Original Assignee: Biocept, Inc.
• Inventors: Tsinberg, Pavel
• Primary Examiner(s): Lundgren; Jeffrey S.
• Application Number:
  US10/922,391
• Publication Number:
  US 20060040274A1
• Time in Patent Office: 1,867 Days
• Field of Search: None
• US Class Current: 435/6.11
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00387 Applications using probes
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/0063 Other, e.g. van der Waals forces, hydrogen bonding
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00641 the porous medium being continuous, e.g. porous oxide substrates
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: G01N 33/54353 with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: G01N 33/54393 Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding
• CPC Class Codes: Y10S 524/916 Hydrogel compositions

Abstract

A method of making a microarray by coating a flat substrate with a polymerizable hydrogel layer which contains anchoring moieties dispersed uniformly therethroughout. Following curing, a continuous layer of uniform thickness is securely attached to the upper surface of the substrate through an array region thereof. A plurality of different probes are then attached to create microspots at distinct spatial locations on the surface of this slab layer by linking the probes to the anchoring moieties in the cured hydrogel. Such anchoring moieties may employ linking systems such as organic chelators, that are activated by copper or some other metal, and complementary pairs such as avidin-biotin.

Claims

1. A microarray for assaying for target material in a biological sample, which microarray comprises:

a solid, flat substrate having an upper surface that is functionalized with organic molecules,

a polymerized hydrogel continuous layer of uniform thickness of at least about 5 microns completely covering an array region of the surface, the layer including anchoring moieties in the form of either organic chelators or streptavidin covalently linked to the hydrogel via isocyanate groups thereof and dispersed uniformly therethroughout, the hydrogel comprising a prepolymer of (a) PEG or PPG or a copolymer thereof of a molecular weight between about 5,000 and about 30,000 daltons and (b) difunctional and/or polyfunctional isocyanates, which prepolymer has active isocyanate groups present in an amount between about 0.1 meq/g and about 1 meq/g and is crosslinked by urea bonds into a porous hydrogel, and

a plurality of different probes attached at distinct spatial locations throughout the at least about 5 micron thick hydrogel layer covering the upper surface to create a plurality of microspots,

the probes being linked to the anchoring moieties in said polymerized hydrogel layer, and regions surrounding each of the microspots being essentially free from probes and resistant to non-specific binding of target materials.