An Improved Immunoperoxidase Technique for Identifying SV40 V and T Antigens by Light Microscopy

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Abstract
Development of the [horseradish] peroxidase reaction for a period of 10 min has consistently resulted in strong nuclear staining of SV40 V [virion] and T [tumor] positive cells [green monkey kidney AH-1 cells, SV40-transformed hamster embryo 2K cells and SV40 transformed hamster embryo SK/B cells]. These cells are easily distinguished from antigen negative cells [hamster embryo line I cells] except for some finely granular cytoplasmic staining present in the latter cells. In addition to its slightly greater sensitivity, the indirect immunoperoxidase method offers the following advantages when compared to the immunofluorescence technique: it allows examination of the preparations under brightfield light microscopy; since the procedure is based on a gentle although highly efficient conjugation reaction, neither the antibody nor the enzyme activity appears to be reduced to any appreciable degree while the conjugate remains stable; the color intensity of the staining reaction is not dependent upon narrow pH changes or upon the ratio of peroxidase to antibody in the conjugation reaction; and since the preparations are permanently mounted, no alterations of the label can occur.

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