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ELISA solid phase: stability and binding characteristics.

A A Ansari, N S Hattikudur, S R Joshi

    Journal of Immunological Methods
    |November 28, 1985
    PubMed
    Summary
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    Storing solid-phase antibodies (SP-Ab) wet in buffer preserves their activity. Drying solid-phase antibodies causes activity loss due to surface denaturation, not desorption.

    Area of Science:

    • Biochemistry
    • Immunology
    • Materials Science

    Background:

    • Solid-phase antibody (SP-Ab) immobilization is crucial for various immunoassays.
    • Understanding SP-Ab stability is essential for assay reliability and shelf-life.

    Purpose of the Study:

    • To investigate the stability of rabbit anti-human chorionic gonadotropin beta subunit antibodies immobilized on polystyrene surfaces.
    • To determine optimal storage conditions for maintaining SP-Ab activity.

    Main Methods:

    • Antibodies were physically adsorbed onto polystyrene surfaces.
    • SP-Ab stability was assessed under various storage conditions (wet/dry, temperature, buffer composition).
    • Radiolabeled antibodies were used to quantify antibody desorption.

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    Main Results:

    • Wet SP-Ab stored in Tris-HCl buffer or ammonium sulphate solution retained activity for up to 95 days at 6°C/room temperature and 53 days at 40°C.
    • Drying led to significant SP-Ab activity loss, which continued during storage.
    • Sucrose or lactose coating provided only partial protection against activity loss upon drying.
    • Antibody desorption was minimal (<1.8%) even after storage in 6 M urea or 0.1% acetic acid, indicating strong binding.
    • Activity loss was attributed to surface denaturation rather than antibody desorption.

    Conclusions:

    • Solid-phase antibodies should be stored wet in a suitable buffer solution.
    • Drying of SP-Ab should be avoided to prevent loss of binding activity.
    • The strong antibody-surface interaction suggests denaturation is the primary cause of activity loss upon drying.