Pretreatment in a high-pressure microwave processor for MIB-1 immunostaining of cytological smears and paraffin tissue sections to visualize the various phases of the mitotic cycle

In many pathology laboratories, both microwave ovens and pressure cookers are used for pretreatment of cytologic smears and paraffin sections to allow MIB-1 staining. For both methods there are two problems. First, the results cannot be used for quantitation because standardization is impossible. Second, the staining results are often suboptimal, resulting in negative staining of cells in the G(1)- and S-phases. When pretreatment is performed in a microwave processor, allowing microwave heating under pressure, precise temperature monitoring becomes possible. In addition, the importance of the pH of the buffer was studied using a test battery series. Optimal staining is achieved at a temperature of 115C, 10 min, pH 6. This method proved to be highly reproducible. Because the immunostaining results are optimal, the various phases of the cell cycle can be defined in the sections and smears. In addition, the perinucleolar staining of the late G(1)-phase is optimally visualized and nuclei of the stable pKi-67 pathway can be identified. Under suboptimal conditions, in particular, the number of cells in the late G(1)-phase are underestimated in the MIB-1 counts.