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An Enzyme-Linked Procedure for the Detection and Estimation of Surface Receptors on Cells¹

From the Department of Microbiology, College of Physicians and Surgeons of Columbia University, New York, New York

Abstract

An enzyme immunoassay procedure has been developed for the visualization and estimation of lymphocyte surface receptors. -Galactosidase (-gal'ase) was covalently linked to sheep anti-rabbit immunoglobulin (SARIg), to ovalbumin (OA), and to adenosine (A). Exposure of rabbit peripheral lymphocytes to SARIg--gal'ase and subsequent incubation with the fluorogenic substrate fluorescein--digalactopyranoside allowed visualization of B cells by fluorescence microscopy. Treatment with each of the above -gal'ase conjugates and incubation with another fluorogenic substrate, 4-methyl--D-umbelliferylgalactopyranoside, made possible the estimation of the various receptors by spectro fluorimetry. Procedures used for enrichment of T and B cell populations could be monitored. Among the findings was that immunoglobulin-bearing cells (presumably B cells) formed rosettes with sheep erythrocytes. The quantitative procedure was used to study lymphocyte population changes during immunization with an A-O A conjugate. An increase in the binding of A--gal'ase, SARIg--gal'ase, and OA--gal'ase by peripheral lymphocytes occurred repeatedly 6 to 8 days after immunization but lasted only a few days despite a continued high circulating antibody titer. These results are consistent with the possibility that immunization induces migration of lymphocytes into the circulation from bone marrow and/or thymus.

Footnotes

¹ This work, which was supported by Research Grant AI-06860 from the National Institutes of Health, is part of a dissertation to be submitted by Deborah J. Cameron in partial fulfillment of the requirements for the Ph.D. degree in the Department of Microbiology, Columbia University.

² Deborah J. Cameron is a Montgomery Maze Fellow.