Protocol

Protein Arrays

Volume 264 of the series Methods in Molecular Biology pp 161-172

A Protein Microarray ELISA for Screening Biological Fluids

  • Susan M. VarnumAffiliated withBiological Sciences Division, Pacific Northwest National Laboratory
  • , Ronald L. WoodburyAffiliated withBiological Sciences Division, Pacific Northwest National Laboratory
  • , Richard C. ZangarAffiliated withBiological Sciences Division, Pacific Northwest National Laboratory

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Abstract

Protein microarrays permit the simultaneous measurement of many proteins in a small sample volume and therefore provide an attractive approach for the quantitative measurement of proteins in biological fluids, including serum. This chapter describes a microarray enzyme-linked immunosorbent assay (ELISA). Capture antibodies are immobilized onto a glass surface; the covalently attached antibodies bind a specific antigen from a sample overlaying the array. A second, biotinylated antibody that recognizes the same antigen as the first antibody, but at a different epitope, is then used for detection. Detection is based on an enzymatic signal-enhancement method known as tyramide signal amplification (TSA). By coupling a microarray-ELISA format with the signal amplification of tyramide deposition, the assay sensitivity is as low as sub-pg/mL.

Key Words

Microarray ELISA tyramide antibody biological fluids serum