Fixation time and microwave oven irradiation affect immunocytochemical p53 detection in formalin-fixed paraffin sections

Maria Grazia Daidone, Elvira Benini, Stefania Rao, Silvana Pilotti, Rosella Silvestrini

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The pathogenetic and prognostic role ascribed by several studies to p53 alterations and the availability of antibodies that recognize p53 protein in paraffin-embedded tissue prompted translational research on the clinical role of p53 accumulation, mainly in breast cancer. However, few studies evaluated the possible interference of fixation time and use of antigen retrieval on p53 detection. In this study, the comparative analysis of immunoreactivity, detected by three p53-specific antibodies (PAb1801, DO7, and CM1), on 87 6- hour formalin-fixed breast carcinomas showed a remarkable agreement in qualitative and quantitative evaluations. The interference of fixation time and microwave oven (MWO) irradiation was analyzed in a subset of 40 randomly selected cases, in which specular specimens were formalin fixed for 6 or 24 hours. Among the 20 cases in which p53 accumulation was detectable after 6 hours of fixation, p53 immunoreactivity disappeared in 14 (70%) after 24- hour fixation without antigen retrieval and was restored in 13 (93%) by MWO irradiation. MWO treatment significantly increased the number of p53- immunoreactive cells compared with those detected after a short-term (6- hour) fixation but did not affect the lack of immunoreactivity of p53- negative cases. These results could have practical implications in the presence of a significant relation between the fraction of p53-positive cells, considered as a continuous variable, and clinical outcome. Moreover, they support the need for a methodologic standardization to guarantee interlaboratory reproducibility of p53 immunostaining and to allow interinstitutional comparison of results.

Original languageEnglish
Pages (from-to)140-144
Number of pages5
JournalApplied Immunohistochemistry
Issue number3
Publication statusPublished - 1998


  • Antigen retrieval
  • Breast cancer
  • Formalin fixation
  • Immunocytochemistry
  • Microwave oven irradiation
  • p53 accumulation

ASJC Scopus subject areas

  • Anatomy
  • Medical Laboratory Technology


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