A monoclonal mouse anti-human IL-2 receptor antibody (anti-Tac) will recognize molecules on the surface of Xenopus laevis immunocytes which specifically bind rIL-2 and are only slightly larger than the human Tac protein.
We have made visible the binding of a mouse monoclonal anti-human interleukin 2 (IL-2) receptor (anti-Tac) antibody on the surface of phytohemagglutinin (PHA)-stimulated Xenopus thymocytes using a colloidal gold-conjugated goat anti-mouse antibody and transmission electron microscopy. No binding was found when a different mouse monoclonal antibody (mAb) of the same isotype and subclass was tested, or when the anti-Tac antibody was omitted from the procedure. After metabolic radiolabeling of the IL-2 receptors with [35S]methionine using PHA-stimulated thymocytes of Xenopus laevis, the South African clawed toad, we show that a concentrated preparation of the mouse anti-human Tac antibody will immunoprecipitate a radiolabeled molecule just slightly larger than 55 kDa. Phorbol dibutyrate (PDB), an effective T cell mitogen, and cyclosporin A, an inhibitor of T cell mitogenesis in this species, are both capable of regulating the expression of this IL-2-binding molecule on Xenopus immunocytes. Here, we use the calcium ionophore A23187 to show that the relationship between IL-2 receptor expression and mitogenesis, which was previously established in X. laevis, is associated with a calcium ion flux. Flow cytometry is used for assaying alterations in epitope expression after binding the lectin-stimulated cells under test with a fluorescence (Fl*) conjugate of the anti-Tac antibody or a control mAb, which is either anti-DNP or anti-keyhole limpet hemocyanin (KLH) in specificity, but of the same mouse isotype and subclass as the anti-IL-2 receptor antibody.
Ruben
L N
LN
Department of Biology, Reed College, Portland, OR 97202-8199.
Langeberg
L
L
Malley
A
A
Clothier
R H
RH
Beadling
C
C
Lee
R
R
Shiigi
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015704:CD4 Antigens / Q000276:immunology*; D002118:Calcium / Q000502:physiology; D007376:Interleukin-2 / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects; D051379:Mice; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D015375:Receptors, Interleukin-2 / Q000276:immunology* / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D013045:Species Specificity; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D014982:Xenopus laevis / Q000276:immunology*
D000911:Antibodies, Monoclonal; D015704:CD4 Antigens; D007376:Interleukin-2; D015375:Receptors, Interleukin-2; D011994:Recombinant Proteins; D015240:Phorbol 12,13-Dibutyrate; D002118:Calcium