Fig. 3

Detection of GSTP1 promoter in SEEMLIS-enriched DNA from prostate cancer CTCs. Circulating tumor cells (CTCs) were enriched by positive selection for EpCAM using ESP. RNA and DNA were extracted from the EpCAM-selected population following live cell, on chip imaging of selected cells. A Gene expression was determined by qPCR for the indicated genes. Raw Ct values were used to create the heat map. Heat map intensity is determined separately for each gene, and comparisons can be made within each column, but not across rows. B DNA extracted from the enriched population was digested with AluI and HhaI restriction enzymes prior to enrichment of methylated DNA by MBD2-MBD. qPCR was performed for GSTP1 and LINE1 using the enriched methylated DNA. MI was calculated by delta Ct relative to a max cycle limit of 45 with all undetected samples (ND) set to a Ct value of 45 for analysis. Optimal threshold for GSTP1 is shown as a dotted line. C MI for GSTP1 and LINE1 is plotted against the number of CTCs (GSTP1) or total number of cells (LINE1). A semi-log nonlinear fit was performed to determine the best fit line. GSTP1 is positively correlated to CTC number with an R² value of 0.94 and p value of 0.0062