A high-throughput assay for the detection of Tyr-phosphorylated proteins in urine of bladder cancer patients.

摘要: Abstract Background Bladder cancer has the peculiarity of shedding neoplastic cells and their components in urine representing a valuable opportunity to detect diagnostic markers. Using semi-quantitative method we previously demonstrated that levels Tyr-phosphorylated proteins (TPPs) are highly increased bladder tissues soluble TPPs can also be detected patient's samples. Although preliminary evaluation showed very promising specificity sensitivity, insufficient accuracy low throughput halted new marker. To overcome this problem developed quantitative methodology with high sensitivity measure urine. Methods The Immobilized Metal Affinity Chromatography (IMAC) was miniaturized 96 well format. Luminescence, visible infrared fluorescence antibody-based detection methods were comparatively evaluated. Results Due abundance evidenced both phosphoprotein enrichment step sensitive required pursue throughput, reproducibility cost containment, which for screening programs, coupled pre-analytical IMAC procedure phases (infrared or chemiluminescence) an automated platform. Conclusions A measuring TPP samples is now available large clinical trial establishment predictive power as General significance assay represents first measurement