Antibody array platform to monitor protein tyrosine phosphorylation in mammalian cells

Protein tyrosine phosphorylation plays a central role in cell-signaling and is a focus of biomedical studies and cancer therapy. However, it is still challenging to identify or characterize the coordinated changes of many candidate proteins of one particular pathway or multiple pathways simultaneously. Antibody array is a recently developed approach applied for differential analysis of multiple protein posttranslational modification events in mammalian cells. It is based on the highly specific recognition between the immobilized antibodies on the array and their specific target proteins in a high-throughput screening format. Here we have described in detail two methods for differential analysis of protein tyrosine phosphorylation in cells by (1) using a single fluorescent protein capture format on membrane array and (2) a competitive protein capture method on glass surface array.