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Related Experiment Videos

Fixation-dependent cytoplasmic false-positive staining with an immunoperoxidase method.

V Bergroth, S Reitamo, Y T Konttinen

    Histochemistry
    |January 1, 1982
    PubMed
    Summary

    Fixation with alcohol or acetone in immunoperoxidase staining can cause false positives in human spleen samples. This nonspecific staining, mainly from the second antibody layer, highlights the need for proper controls in diagnostic tests.

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    Area of Science:

    • Histopathology
    • Immunocytochemistry
    • Biomedical Research

    Background:

    • The unlabeled immunoperoxidase (PAP) method is widely used for tissue staining.
    • Nonspecific staining can lead to misinterpretation of results in diagnostic pathology.

    Purpose of the Study:

    • To investigate fixation-dependent nonspecific staining in the PAP method.
    • To identify the cause of false-positive staining in human spleen sections.

    Main Methods:

    • Human spleen sections were embedded in paraffin.
    • Sections were fixed using various fixatives, including acetone, alcoholic formalin, and absolute ethanol.
    • The primary antiserum was omitted or replaced with normal rabbit serum to assess nonspecific staining.

    Main Results:

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    • Strong cytoplasmic staining was observed in polymorphonuclear leucocytes and macrophages after fixation with acetone, alcoholic formalin, and absolute ethanol.
    • The observed staining was primarily attributed to the second layer of the PAP method.
    • Nonspecific protein-immunoglobulin interaction due to alcoholic or acetone fixation is the likely cause.

    Conclusions:

    • Alcoholic and acetone fixatives can induce significant nonspecific staining in the PAP method.
    • The findings underscore the critical importance of including appropriate controls in all PAP staining procedures.
    • Implementing proper controls is essential to prevent false-positive interpretations in histopathological diagnoses.