Microinjection of macromolecules into normal murine lymphocytes by cell fusion technique. I. Quantitative microinjection of antibodies into normal splenic lymphocytes.
Human erythrocyte ghosts loaded with various kinds of protein molecules were fused with mouse splenic lymphocytes by means of polyethylene glycol supplemented with poly-L-arginine and dimethylsulfoxide. This fusion method made quantitative microinjection of IgG and other proteins into intact lymphocytes possible. The injection itself did not alter cell viability, and lymphocytes given protein molecules retained intact response activity when they were stimulated with mitogens. Rabbit anticyclic AMP was purified by affinity chromatography and injected into lymphocytes. Antibody activity in the cell lysates was measured by using 125I-labeled cyclic AMP as an antigen, and it was shown that antibody molecules were quantitatively injected and immunologically active in the cells. Antigen binding activity of anti-cyclic AMP antibodies in the nonstimulated lymphocytes was stable and intact even 24 hr after microinjection, whereas the activity rapidly decreased in mitogen-stimulated lymphocytes, indicating that some immunologic or enzymatic mechanisms for inactivating antibodies were induced in mitogen-stimulated cells. Furthermore, microinjection of anti-cyclic AMP markedly enhanced the proliferative responses of lymphocytes to mitogens such as Con A or LPS and reversed the effect of a known elevator of intracellular cyclic AMP. These observations have implications for the role of cyclic AMP in early lymphocyte activation events.
D000818:Animals; D000906:Antibodies / Q000502:physiology*; D002459:Cell Fusion; D000242:Cyclic AMP / Q000276:immunology; D004247:DNA / Q000096:biosynthesis; D004910:Erythrocyte Membrane / Q000378:metabolism; D005260:Female; D006801:Humans; D007158:Immunologic Techniques; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000276:immunology* / Q000502:physiology; D046911:Macromolecular Substances; D008297:Male; D051379:Mice; D008845:Microinjections; D011092:Polyethylene Glycols / Q000494:pharmacology; D011817:Rabbits; D013154:Spleen / Q000166:cytology
D000906:Antibodies; D046911:Macromolecular Substances; D011092:Polyethylene Glycols; D004247:DNA; D000242:Cyclic AMP