Preparing glass slides and coverslips for in situ hybridization

INTRODUCTIONPrecleaned glass slides are of high enough quality for both in situ and immunohistochemical techniques. However, for in situ hybridization, the slides need to be treated with diethyl pyrocarbonate (DEPC) so that any RNase attached to them is destroyed. The slides also need to be coated with 3-triethoxysilylpropylamine (TESPA) or poly-L-lysine so that the sections adhere tightly and do not detach during subsequent extensive washing procedures. This protocol describes techniques for coating slides with TESPA and poly-L-lysine. There are advantages and disadvantages to each coating method. TESPA-treated slides can be stored for a long time, but the sections do not adhere tightly until after drying. Poly-L-lysine-coated slides need to be made fresh, but the sections adhere immediately on contact with the surface. This protocol also describes how to prepare coverslips for in situ hybrization by coating them in a siliconizing solution.