Methods Mol. Biol. 723 (-) 29-36 [2011-03-04; online 2011-03-04]
Alongside the increasing availability of affinity reagents, antibody microarrays have been developed to become a powerful tool to screen for target proteins in complex samples. Besides multiplexed sandwich immunoassays, the application of directly applying labeled sample onto arrays with immobilized capture reagents offers an approach to facilitate a systematic, high-throughput analysis of body fluids such as serum or plasma. An alternative to commonly used planar arrays has become available in form of a system based on color-coded beads for the creation of antibody arrays in suspension. The assay procedure offers an uncomplicated option to screen larger numbers of serum or plasma samples with variable sets of capture reagents. In addition, the established procedure of whole sample biotinylation circumvents the purification steps, which are generally required to remove excess labeling substance. We have shown that this assay system allows detecting proteins down into lower pico-molar and higher picogram per milliliter levels with dynamic ranges over three orders of magnitude. Presently, this workflow enables the profiling of 384 clinical samples for up to 100 proteins per assay.