Development of 3 Multi-PCR Based Microsatellite Instability Analysis (MSA) for Bladder Cancer Detection

Several studies have shown that microsatellite changes can be profiled in urine for the detection of bladder cancer. Use of microsatellite analysis (MSA) for bladder cancer detection requires a comprehensive analysis of up to 15 to 20 markers, which were based on amplification and interpretations of many individual MSA markers and can be technically challenging. Here, in a way to develop fast, more efficient, standardized and less costly MSA for detection of bladder cancer, we have developed 3 multiplex PCR based MSA assay, all of which were analyzed by genetic analyzer. First, we have selected 16 MSA markers based on 9 selected publications. Based on samples from Johns Hopkins University (JHU Sample, first set sample), we have attempted to develop MSA assay based on doublet, 2 tube based multiplex PCR combined with singlet, 1 tube based single plex PCR. While this assay was initially successful, we have encountered a reproducibility issue and we then developed MSA based on triplet, 3 tube based multiplex PCR (Triplet MSA assay). From second set samples (6 cancer patients’ and 14 normal individuals’ sample), our Triplet Assay with 15 MSA markers correctly predicted all of 6/6 cancer samples to be cancerous sample and 14/14 samples from normal individuals as normal samples. This result suggests that our Triplet MSA Assay combined with genetic analyzer is a potentially time and cost-effective genetic assay for bladder cancer detection and can be used potentially as a dependable assay in patient care.