High-Resolution Silver Staining-Based PCR-Single Strand Conformation Polymorphism (SSCP) Analysis

In this chapter, we describe a high-resolution non-radioactive PCR-SSCP method which utilizes a polyacrylamide-agarose composite gel and background-free silver staining. Addition of agarose into the acrylamide solution produces a strong and flexible gel for SSCP analysis, while treatment of the gel with sodium thiosulfate prior to and after exposure to silver salts results in background-free staining. Together, both modifications improve the resolution of silver staining-based PCR-SSCP analysis and increase the sensitivity of mutation detection to a level similar to that of radioactive labeling-based methods.