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Fluorescent Immunohistochemistry and In Situ Hybridization Analysis of Mouse Pancreas Using Low-power Antigen-retrieval Technique

Shundi Ge, Gay M. Crooks, George McNamara and Xiuli Wang

Division of Research Immunology/Bone Marrow Transplantation, Childrens Hospital, Los Angeles, California (SG,GMC,XW), and Division of Cancer Immunotherapeutics and Tumor Immunology, City of Hope National Medical Center, Duarte, California (GM)

Correspondence to: Gay M. Crooks, MD, Division of Research Immunology/BMT, Childrens Hospital Los Angeles, 4650 Sunset Blvd. MS #62, Los Angeles, CA 90027. E-mail: gcrooks{at}chla.usc.edu

To facilitate the immunological reaction of antibodies with antigens in fixed tissues, it is necessary to unmask or retrieve the antigens through pretreatment of the specimens. However, adjustment of heating-induced antigen retrieval is always required for different tissues and antigens. In this study, by using a low-power antigen-retrieval technique with appropriate dilution of antibodies, we successfully immunostained key antigens in pancreas such as insulin, PDX-1, glucagon, cytokeratin, and CD31, which have previously presented a particular challenge for investigators because of the rapid autodigestion and high nonspecific antibody binding in this tissue. Satisfactory results were obtained when immunohistochemistry and fluorescence in situ hybridization analysis were combined in the same slides. (J Histochem Cytochem 54:843–847, 2006)

Key Words: heating-induced antigen retrieval • low-power antigen-retrieval technique • fluorescence immunohistochemistry

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