Possible role of tissue‐bound calcium ions in citrate‐mediated high‐temperature antigen retrieval

J. M. Morgan, H. Navabi, K. W. Schmid, B. Jasani

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146 Citations (Scopus)


High‐temperature preheating of sections in the presence of a salt (e.g., citrate) or a protein denaturant (e.g., urea) solution has been shown recently to provide a reliable alternative to tissue proteolysis for antigen retrieval from formaldehyde‐fixed, paraffin‐embedded tissues. However, the underlying mechanism of action of this form of pretreatment remains highly speculative. In this study, we show that calcium chelating agents such EDTA and EGTA are more effective than citrate in the retrieval of a citrate‐sensitive nuclear antigen, Ki‐67. Also, sodium carbonate and another calcium precipitating agent are both able to effect antigen retrieval at high temperatures. The overall data therefore suggest that either the chelation or the precipitation of tissue‐bound calcium ions, and perhaps also other divalent metal cations, is a critical step in salt‐mediated antigen retrieval. As a corollary, it is suggested that tight complexing of calcium ions or other divalent metal cations with proteins during formaldehyde tissue fixation is responsible for the masking of certain antigens.

Original languageEnglish
Pages (from-to)301-307
Number of pages7
JournalThe Journal of pathology
Issue number4
Publication statusPublished - Dec 1994



  • High temperature
  • Ki‐67
  • antigen retrieval
  • autoclave
  • calcium ion chelation
  • formaldehyde fixation
  • immunohistochemistry
  • microwave

ASJC Scopus subject areas

  • Pathology and Forensic Medicine

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