Microwave antigen retrieval in immunocytochemistry: a study of 80 antibodies

AIMS--To evaluate the effect of microwave irradiation on the staining quality of a range of commonly used primary antibodies in archival, formalin fixed, paraffin wax embedded material, with emphasis on antibodies that have previously worked successfully only on frozen tissue. METHODS--Immunocytochemistry (streptavidin-biotin complex technique) was performed on histological sections of a range of normal and pathological tissues, after varying treatment with microwave irradiation. The staining quality of each antibody was compared with that achieved without prior treatment of the sections or after enzyme predigestion. RESULTS--Microwave irradiation permitted successful immunostaining with 20 antibodies that stained only frozen tissues before. The staining characteristics of 21 antibodies that were already known to stain formalin fixed, paraffin wax embedded material were improved. Another 39 antibodies did not show enhanced staining with microwave irradiation. The method preserves tissue morphology and produces more consistent staining than that achieved by enzyme predigestion with many antibodies. Microwave irradiation may also allow some primary antibodies to be used at higher working dilutions. The citrate buffer used in this study avoids the necessity of exposure to heavy metal salts. CONCLUSIONS--Microwave antigen retrieval represents an important technical advance within immunocytochemistry that will greatly increase the range of antibodies which can be used to study formalin fixed, paraffin wax embedded tissues.