Effective radioiodination by lactoperoxidase and solubilisation of cell-surface proteins of cultured murine T lymphoma cells

Conditions for cell-surface radioiodination by lactoperoxidase developed previously for lymphocytes were found to be suboptimal for iodination of cells of different cultured mouse T lymphoma lines. The efficiency of iodination of these cells was greatly increased by adjusting the total iodide concentration to 20 μM in the reaction mixture and reinitiating the enzyme-catalyzed reaction by repeated additions of lactoperoxidase and hydrogen peroxide at regular time intervals. By this procedure about 3 x 105 atoms of [125I] iodide/lymphoma cell could be incorporated without loss of cell viability. Techniques for solubilisation of radioiodinated cell-surface proteins of T lymphoma cells by metabolic release and by extraction with non-ionic detergent NP40, NP40-urea or urea-acetic acid were studied and compared. Only extraction with NP40-urea was sufficient to obtain 90-95% of cell-associated radioactivity in solution. Immunoglobulin was isolated and quantitated by specific coprecipitation from surface proteins of the cells of one lymphoma line (WEHI-22). It was found that the percentage of immunoglobulin in solubilised labelled cell-surface proteins was strongly dependent on the solubilisation technique used