Journal oflmmunologicalMethods, 65 (1983) 369-372 369 Elsevier JIM02873 Cloning of Lymphocytes from Whole Blood by Limiting Dilution Kevin J. Trainor and Alexander A. Morley Department of Haematology, Flinders Medical Centre, Bedford Park Adelaide, South Australia 5042, Australia (Received 25 March 1983, accepted9 August 1983) A simple whole blood limiting dilution technique was developed to clone human lymphocytes non-specifically.The geometricmean frequencyof clone formingcells in 13 normal individuals was found to be 31.1%. Compared with measurement of proliferation in mass suspension culture, cloningprovides a quantitative and easily interpretable endpoint for a variety of lymphocyte studies and its use in measurement of radiation sensitivity is presented as an example. Key words: lymphocyte cloning - limiting dilution cloning - cloning frequency Introduction The lymphocyte response to mitogens is commonly used for assessment of cellular immune function or as an endpoint for quantitation of a variety of experimental manipulations. The mitogen response in suspension culture is, however, a complex phenomenon which involves interactions between several populations of cells and concomitant death and proliferation of cells. Owing to this, interpretation of any quantitative response in terms of the response of a particular cell type is difficult and any quantitative conclusion can only be approximate. Cloning techniques are now widely used for culturing lymphocytes, and much interest has been devoted to the cloning of immunologically defined subpopulations. A non-specific cloning technique of high efficiency would be of great value in providing a quantitative and easily interpreted endpoint for a variety of studies. Recently we reported such as cloning technique for separated lymphocytes (Morley et al., 1983) and in this paper we report a simple high efficiency cloning technique for whole blood. To illustrate the use of this technique as an endpoint, we describe the results of measurement of sensitivity of lymphocytes to X-irradiation. 0022-1759/83/$03.00 © 1983 Elsevier SciencePublishers B.V.