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New immunolatex spheres: visual markers of antigens on lymphocytes for scanning electron microscopy

Molday, Robert S. and Dreyer, William J. and Rembaum, Alan and Yen, S. P. S. (1975) New immunolatex spheres: visual markers of antigens on lymphocytes for scanning electron microscopy. Journal of Cell Biology, 64 (1). pp. 75-88. ISSN 0021-9525. http://resolver.caltech.edu/CaltechAUTHORS:20120802-083522605

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Abstract

New immunochemical reagents consisting of antibodies bound to small latex spheres were used as visual markers for the detection and localization of cell surface antigens by scanning electron microscopy. Cross-linked latex spheres of various sizes from 300 to 3,4000 A in diameter were synthesized by aqueous emulsion copolymerization of methacrylate derivatives containing hydroxyl and carboxyl functional groups. Proteins and other molecules containing primary amino groups were covalently bonded to the acrylic spheres under a variety of mild conditions by the aqueous carbodiimide, cyanogen bromide, and glutaraldehyde methods. For use in the indirect immunochemical-labeling technique, goat antibodies directed against rabbit immunoglobulins were bonded to the spheres. These immunolatex reagents were shown to bind only to cells (red blood and lymphocytes) which had previously been sensitized with rabbit antibodies against cell surface antigens. Mouse spleen lymphocytes with exposed immunoglobulins on their surface (B cells) were labeled with these spheres and distinguished from unlabeled or T lymphocytes by scanning electron microscopy. The distribution of Ig receptors on lymphocytes was also studied using the spheres as visual markers. When lymphocytes were fixed with glutaraldehyde and subsequently labeled with the immunolatex reagents, a random distribution was observed by scanning electron microscopy; a patchy distribution was observed when unfixed lymphocytes were used. These results are consistent with studies using ferritin-labeled antibodies (S. De Petris and M. Raff. 1973. Nature [Lond.]. 241:257.) and support the view that Ig receptors on lymphocytes undergo translational diffusion. In addition to serving as visual markers for scanning electron microscopy, these latex spheres tagged with fluorescent or radioactive molecules have applications as highly sensitive markers for fluorescent microscopy and as reagents for quantitative studies of cell surface antigens and other receptors.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1083/jcb.64.1.75 DOIUNSPECIFIED
http://jcb.rupress.org/content/64/1/75.abstractPublisherUNSPECIFIED
Additional Information:© 1975 The Rockefeller University Press. Received for publication I July 1974, and in revised form 19 September 1974. This work was supported by grants from the President's Fund of the California Institute of Technology, U. S. Public Health Service grants GM 06965 and CA 15926, and National Institutes of Health contract no. NCI-CB-43875. R. S. Molday was supported by an American Cancer Society postdoctoral grant.
Funders:
Funding AgencyGrant Number
Caltech President’s FundUNSPECIFIED
Public Health Service grantGM 06965
Public Health Service grantCA 15926
NIHNCI-CB-43875
American Cancer Society postdoctoral grantUNSPECIFIED
Record Number:CaltechAUTHORS:20120802-083522605
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20120802-083522605
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:32863
Collection:CaltechAUTHORS
Deposited By: Jason Perez
Deposited On:02 Aug 2012 18:21
Last Modified:26 Dec 2012 15:47

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