Use of porous membranes modified with polyelectrolyte multilayers as substrates for protein arrays with low nonspecific adsorption.
Coating of substrates with polyelectrolyte multilayers terminated with poly(acrylic acid) (PAA) followed by activation of the free -COOH groups of PAA provides a surface that readily reacts with amine groups to allow covalent immobilization of antibodies. The use of this procedure to prepare arrays of antibodies in porous alumina supports facilitates construction of a flow-through system for analysis of fluorescently labeled antigens. Detection limits in the analysis of Cy5-labeled IgG are 0.02 ng/mL because of the high surface area of the alumina membrane, and the minimal diameter of the substrate pores results in binding limited by kinetics, not mass transport. Moreover, PAA-terminated films resist nonspecific protein adsorption, so blocking of antibody arrays with bovine serum albumin is not necessary. These microarrays are capable of effective analysis in 10% fetal bovine serum.
Dai
Jinhua
J
Department of Chemistry, Michigan State University, East Lansing, Michigan 48824, USA.
Baker
Gregory L
GL
Bruening
Merlin L
ML
D000180:Acrylic Resins / Q000737:chemistry; D000327:Adsorption; D000537:Aluminum Oxide / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D000937:Antigen-Antibody Reactions; D001672:Biocompatible Materials / Q000737:chemistry*; D002417:Cattle; D004573:Electrolytes / Q000737:chemistry* / Q000145:classification / Q000494:pharmacology; D005456:Fluorescent Dyes; D007118:Immunoassay; D007074:Immunoglobulin G / Q000737:chemistry*; D008567:Membranes, Artificial; D051379:Mice; D016062:Porosity; D040081:Protein Array Analysis; D011506:Proteins / Q000737:chemistry*; D011817:Rabbits; D051381:Rats; D012710:Serum Albumin, Bovine; D013499:Surface Properties
D000180:Acrylic Resins; D001672:Biocompatible Materials; D004573:Electrolytes; D005456:Fluorescent Dyes; D007074:Immunoglobulin G; D008567:Membranes, Artificial; D011506:Proteins; D012710:Serum Albumin, Bovine; C006903:carbopol 940; D000537:Aluminum Oxide