Inhibition of plaque-forming cells with anti-idotope or hapten: Variation due to hapten density on indicator red cells

Cronkhite, R; Černý, Jan; DeLisi, Charles

doi:10.1016/0022-1759(84)90141-8

Cited by 3 publications

(4 citation statements)

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“…Antigens such as polyvinyl pyrrolidone, pneumococcal polysaccharides or haptens such as arsonate or 4-hydroxy 5-iodo 3-nitrophenyl, phosphoryl choline have also been conjugated to SRBC and used in PFC assays (Conger et al, 1983;Cronkhite et al, 1984;Garg et al, 1994;Goud et al, 1992;Swenson et al, 1997). 6.…”

Section: Methodsmentioning

confidence: 99%

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Assays for B Lymphocyte Function

Bondada

Robertson

2003

CP in Immunology

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This unit describes the antigenic stimulation of in vitro antibody production by B cells and the subsequent measurement of secreted antibodies. The first basic protocol is a generalized system for inducing in vitro antibody production and can accommodate various types of antigens under study. Secreted antibodies can then be measured with an enzyme-linked immunosorbent assay (ELISA) or other soluble-antibody detection systems. Alternatively, the number of antibody-producing cells can be quantified by plaque-forming cell (PFC) assays presented in this unit: the Cunningham-Szenberg and the Jerne-Nordin techniques. Both methods employ specially prepared slide chambers, described here, in which the antibody-producing B cells are mixed with complement and indicator sheep red blood cells (SRBC), or with trinitrophenol-modified SRBC (TNP-SRBC), with subsequent lysis and counting of plaques. Because IgM antibodies fix complement efficiently, whereas IgG and IgA antibodies do not, unmodified PFC assays measure only IgM antibodies. The assay can be modified, however, to measure all classes of antibodies or to enumerate total immunoglobulin-secreting B cells, as described in alternate protocols. Yet another method of measuring the number of antibody-producing B cells (in a class-specific fashion) is to use the ELISPOT technique described in UNIT 7.14. The resting B cells used in these procedures are prepared as described in the final support protocols for Percoll gradient centrifugation.

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Section: Methodsmentioning

confidence: 99%

“…Generally, trinitrophenyl groups have been used as haptens in many systems since the PFC assays are well suited for this hapten. Some of the other haptens that can be used are nitroiodophenyl (NIP), nitrophenyl (NP), phosphoryl choline, and arsonate (Conger et al, 1983;Cronkhite et al, 1984;Garg et al, 1994;Goud et al, 1992;Swenson et al, 1997).…”

Section: 818mentioning

confidence: 99%

Assays for B Lymphocyte Function

Bondada

Robertson

2003

CP in Immunology

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“…Plaques developed in the presence of HBSS represented the total number of direct PFC. The addition of phosphorylcholine chloride at M [27] revealed that the proportion of PC-specific, hapten-inhibitable PFC was 2 95% in all experiments. The PFC response in vitro was calculated per culture well (mean f SD from triplicate assays) based on the number of replica cultures pooled.…”

Section: Antibody Pfc Response Assaymentioning

confidence: 97%

“…Sheep red blood cells (Colorado Serum Co., Denver, CO) were covalently coupled to p-diazonium phenylphosphorylcholine as described [27]. Lymphocytes from replica culture wells (8 to 12) were pooled, resuspended in Hanks' balanced salt solution (HBSS) (Microbiological Associates, Bethesda, MD) and assayed for PFC in triplicates in Cunningham-Szenberg chambers constructed from microscopic slides [28].…”

Section: Antibody Pfc Response Assaymentioning

confidence: 99%

Restricted idiotypic profile of anti‐phosphorylcholine antibodies induced by carrier‐specific helper T cell clones

et al. 1989

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The primary antibody response of C57BL/6 mice to phosphorylcholine (PC) is dominated by immunoglobulins of the T15 idiotype family. Using monoclonal antibodies that define individual idiotopes (Id) of T15, it has been shown that the antibody plaque-forming cells (PFC) arise from heterogeneous population of B cells that may express some but not all T15 Id. In the present study, we tested the ability of antigen (keyhole limpet hemocyanin, KLH)-specific CD4+ T cell clones (Th) to provide help for various T15 B cell subsets in response to PC-KLH in vitro. Four independently derived, KLH-specific Th clones were found to provide quantitative help (80-120 PFC/culture of 10(6) B cells) which was comparable to KLH-primed, unselected T cells. However, the response in the presence of the cloned T cells was idiotopically restricted: T15 idiotopes B24-44 and B36-75 were expressed on 60% to 90% PFC, whereas Id AB1-2 and B36-82 were not present (0%-20% PFC). All of these Id were highly expressed in cultures with KLH-primed, unselected splenic T cells. Evidently, the cloned Th cells provided help to only a subset of idiotypically distinct B cells. Bulk spleen T cells from unprimed donors by themselves did not provide any help for the primary response to PC-KLH. However, when these cells were added to the cultures containing B cells and KLH-specific Th clones, the repertoire of the response was fully restored, including the AB1-2/B36-82 Id (50%-80% PFC). These results suggest the existence of an auxiliary helper mechanism that may be required for activation of some B cells in addition to the antigen-specific Th cells.

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Fast plaque sizing and some applications of the technique in antibody feedback experiments

Dresser¹

1990

Journal of Immunological Methods

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Inhibition of plaque-forming cells with anti-idotope or hapten: Variation due to hapten density on indicator red cells

Cited by 3 publications

References 8 publications

Assays for B Lymphocyte Function

Assays for B Lymphocyte Function

Restricted idiotypic profile of anti‐phosphorylcholine antibodies induced by carrier‐specific helper T cell clones

Fast plaque sizing and some applications of the technique in antibody feedback experiments

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