NDLI: A high-efficiency cloning system for single hapten-specific B lymphocytes that is suitable for assay of putative growth and differentiation factors.

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A high-efficiency cloning system for single hapten-specific B lymphocytes that is suitable for assay of putative growth and differentiation factors.

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A high-efficiency cloning system for single hapten-specific B lymphocytes that is suitable for assay of putative growth and differentiation factors.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Nossal, G. J. Pike, B. L.
Abstract	Fluorescein (FLU)-specific murine splenic B lymphocytes from nonimmunized adult mice were prepared by the hapten-gelatin fractionation technique and cultured singly or in very small numbers in 10-microliters culture wells. Growth and differentiation to antibody-secreting status were promoted by polymeric FLU-conjugated antigens with or without added T-lymphocyte-derived conditioned media or purified cytokines. In some cultures, 3T3 fibroblasts or CBA/N thymocytes provided a source of filler cells. Anti-FLU antibody formation was detected by a sensitive enzyme-linked immunosorbent assay (ELISA). With an optimal number (around 300) of 3T3 cells per well, up to 77% of the B cells could be induced to produce detectable antibody. The ELISA permitted detection of antibody formation in essentially all wells where B-cell proliferation occurred, and it was more efficient in detecting antibody-forming clones than the hemolytic plaque assay, whether filler cells were present or not. When 10 B cells rather than 1 were included per well, the ELISA, detecting absorbance in standard fashion, provided a useful method for assessment of B-cell growth- and differentiation-promoting factors (BGDF). It was found that 3T3 cells gave less background stimulation than thymus cells, permitting the detection of as little as 1/100th as much BGDF as with thymocytes, thus offering a dynamic range of up to 30 between control absorbance in the absence of factors and the optimal factor level. Use of 3T3 cells also avoids a potential lymphokine cascade. The system has confirmed that interleukin-2 acts as a BGDF, but it has failed to establish an effect of interferon-gamma on B cells. It has also shown the inactivity of a variety of hemopoietic growth factors on B lymphocytes. This system thus promises to be a useful tool in the further analysis of B-lymphocyte activation.
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	10
Volume Number	82
Language	English
Publisher	National Academy of Sciences
Publisher Date	1985-06-01
Publisher Place	United States
Access Restriction	Open
Subject Keyword	B-Lymphocytes Immunology Growth Substances Lymphokines Animals Antibody Formation Antibody-Producing Cells Ultrastructure Cytology Biological Assay Cell Differentiation Cell Division Cells, Cultured Enzyme-Linked Immunosorbent Assay Fibroblasts Pharmacology Haptens Hemolytic Plaque Technique Interleukin-4 Mice Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S. Multidisciplinary
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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