Perforin, a pore-forming protein detectable by monoclonal antibodies, is a functional marker for killer cells

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Abstract
Perforin Is one of the important cytolytlc factors in cytotoxlc T lymphocytes (CTL) and natural killer (NK) cells. In this paper, we report rat mAbs against mouse perforln established by immunization with a recomblnant mouse perforln fragment. These mAbs reacted with purified mouse perforln prepared from cytoplasmlc granules of an NK-IIke cell line In ELISA and Western blot analysis. However, none of these mAbs blocked the hemolytic activity of mouse perforln or absorbed It when fixed in the solid phase. These results indicate that all of these mAbs react with denatured but not with native mouse perforln. By using a combination of the mAbs, we established a sandwich ELISA, for quantltatlng the cellular contents of perforin. These mAbs were also useful for Immunohlstochemical staining analysis, and perforln was detected in the cytoplasmlc granules of CTL and NK cell lines. Perforln was also detected In a minor population of lymphocytes of the spleen, liver, and lymph node. In normal spleen cells of 5- to 8-week-old mice, 12–15% of aslalo GM1+ cells and 7–21% of CD8+ T cells were perforln-positive, but CD4+ T cells, B cells, and macrophages were totally negative. These data clearly show that perforln Is expressed in cells of a cytotoxlc character In normal mice, In the same way as in primed mice.

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