A simple immunomagnetic bead-based technique for the detection of surface molecules capable of inducing T cell functional polarisation.
Activated T cells can release lymphokines selectively towards the site of contact with the target cell. In this way the specificity of the target-effector cell interaction can be maintained in spite of signalling being mediated by soluble factors (Mosmann, 1988, Immunol. Today 9, 306). However, this polarised phenotype is not expressed in resting T cells; rather it appears to be induced in the first minutes following T cell activation. In order to analyse single molecules for their ability to induce T cell polarisation, we devised a technique based on targeting different T cell surface molecules with specific antibodies immobilised on to immunomagnetic beads. The polarised phenotype was determined from observation of the microtubule organising centre being oriented towards the site of interaction with the bead. When applied to T cell lines, the technique permitted the classification of CD3 as a polarisation-inducing molecule, while no polarisation was found when targeting CD2, CD6 and CD8 molecules. This technique has a number of potential applications since it can, in principle, be applied to any cell surface molecule or cell type. Technical details and the sensitivity of the procedure are discussed.
Rubbi
C P
CP
Department of Biology, University of Essex, Colchester, UK.
Rickwood
D
D
D015703:Antigens, CD / Q000032:analysis; D000954:Antigens, Surface / Q000032:analysis* / Q000502:physiology*; D002448:Cell Adhesion / Q000276:immunology; D016764:Cell Polarity / Q000276:immunology*; D005898:Glass; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D019476:Insect Proteins; D015459:Leukemia-Lymphoma, Adult T-Cell; D008213:Lymphocyte Activation / Q000187:drug effects; D008869:Microtubule-Associated Proteins / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology*; D014407:Tumor Cells, Cultured
D015703:Antigens, CD; D000954:Antigens, Surface; D019476:Insect Proteins; D008869:Microtubule-Associated Proteins; C054628:microtubule-organizing center-associated polypeptide, insect