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Rhodamine immunohistofluorescence applied to plant tissue.

S J Hapner, K D Hapner

    The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society
    |June 1, 1978
    PubMed
    Summary
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    Sanifoin plant tissues autofluoresce brightly under blue light, mimicking fluorescein. Using rhodamine with green light avoids this autofluorescence, improving immunohistochemical staining in plants.

    Area of Science:

    • Plant Biology
    • Biochemistry
    • Microscopy

    Background:

    • Autofluorescence in plant tissues can interfere with immunohistochemical staining.
    • Sanifoin (Onobrychis viciifolia, Scop.) root and seed tissues exhibit strong autofluorescence under blue light (495 nm).
    • This autofluorescence is spectrally similar to fluorescein, a common immunohistochemical fluorochrome.

    Purpose of the Study:

    • To evaluate the utility of rhodamine isothiocyanate for immunohistochemical localization in plant tissues.
    • To identify a fluorochrome that minimizes interference from plant autofluorescence.

    Main Methods:

    • Excitation of sanifoin tissue with blue light (495 nm) to observe autofluorescence.
    • Coupling rhodamine isothiocyanate to immunoglobulin.

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  • Excitation of rhodamine-coupled immunoglobulin with green light (546 nm) and observation with a 580 nm barrier filter.

    • Main Results:

    • Sanifoin root and seed tissues show bright autofluorescence under blue light, comparable to fluorescein.
    • Rhodamine isothiocyanate exhibits reddish-orange fluorescence (emission max 590 nm) when excited at 546 nm.
    • Plant tissues display minimal autofluorescence when excited at 546 nm and viewed with a 580 nm barrier filter.

    Conclusions:

    • Rhodamine isothiocyanate is a suitable fluorochrome for immunohistochemical staining in plant tissues.
    • Utilizing rhodamine with green light excitation (546 nm) effectively circumvents autofluorescence issues in plant samples.
    • This approach enhances the reliability and interpretability of immunohistochemical staining in plant research.