Detection of islet cell specificity of monoclonal islet cell surface antibodies by means of double-staining immunofluorescence

We have generated monoclonal antibodies (mcab) reactive with islet cell surface antigens. 10 different mcab were characterized regarding their islet cell binding specificity by means of a modified double immunofluorescence test. At this assay, the monoclonal islet cell surface antibodies were visualized on rat islet cells by indirect immunofluorescence with fluorescein isothiocyanate-labelled antibodies against mouse immunoglobulin. The alpha and beta cell specificity was determined by indirect immunofluorescence using anti-glucagon or anti-insulin serum and a tetramethyl rhodamine isothiocyanate-labelled second antibody. The target islet cell suspension used contained 61% beta and 23% alpha cells. The monoclonal antibody K28D6 preferentially reacted with alpha cells. The binding of K29aC6 and K56aF3 indicates a high specificity against beta cells. The remaining 7 antibodies were reactive with alpha as well as with beta cells.