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Methodological aspects of the immunostaining of proliferating cell nuclear antigen (PCNA) in cytospin preparations of MCF‐7 cell line
Author(s) -
Pelosi Giuseppe,
Bresaola Enrica,
Menacherry Mary J.,
Manfrin Erminia,
Lannucci Antonio
Publication year - 1994
Publication title -
diagnostic cytopathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.417
H-Index - 65
eISSN - 1097-0339
pISSN - 8755-1039
DOI - 10.1002/dc.2840100120
Subject(s) - proliferating cell nuclear antigen , fixative , immunostaining , immunoperoxidase , immunohistochemistry , alkaline phosphatase , microbiology and biotechnology , pathology , fixation (population genetics) , antigen , staining , medicine , biology , antibody , immunology , monoclonal antibody , biochemistry , enzyme , gene
Cytospins of a human breast cancer cell line (MCF‐7) were studied for the expression of PCNA, a cell cycle‐related protein, using a variety of fixation and immunostaining procedures. The best fixative for PCNA was found to be buffered formaldehyde solution at 4°C followed by methanol at 20°C, whereas alcoholic fixatives decreased greatly the PCNA immunoreactivity. Air‐drying procedures of cytospins prior to and after fixation greatly undermined the PCNA immunostaining. A modified immunoperoxidase method provided a stronger staining of the PCNA‐reactive cells than the alkaline phosphatase anti‐alkaline phosphatase (APAAP) technique. PCNA immunoreactivity could be maintained up to 2 mo, putting slides in methanol at − 2°C. In conclusion, our report indicates that PCNA is a labile antigen, which may critically be affected by temperature and air‐drying procedures. © 1994 Wiley‐Liss, Inc.