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pubmed:abstractText |
To raise monoclonal antibodies that specifically recognize the heavy chain variable region of MOPC141 myeloma protein (VH141), which belongs to VHQ52 family, rats were immunized with Fd'-conjugated keyhole limpet haemocyanin (KLH) (Fd': Fd' fragments of MOPC141), and the spleen cells were fused with mouse myeloma cells. The resulting 900 hybridomas were screened for antibody activity against Fd'1 fragments having no constant H-chain sequences, which were prepared by cleavage of the Fd' fragments with cyanogen bromide, and two monoclonal antibodies, designated 3-2-7h and 3-5-6f, were obtained. Radioimmunoassay inhibition test showed that the two monoclonal antibodies specifically recognized the VH141, but each was directed to a different determinant on the VH141. When the functional VH gene of Abelson virus-transformed mu-producing pre-B cells, which could be strongly stained with 3-5-6f monoclonal antibody, was cloned and sequenced, the VH gene was closely relate to that of MOPC141 (88% and 94% homology at amino acid and DNA level, respectively). Taken together, the results indicated that 3-2-7h had high specificity only for the VH141, whereas 3-5-6f specifically reacted not only with the VH141 but also with the VH region closely related to that of MOPC141, and that both the monoclonal anti-VH141 antibodies were specific for a limited range of VH regions within the VHQ52 family rather than being VHQ52 family specific. These monoclonal anti-VH141 antibodies should be very useful to determine at a single cell level by immunofluorescence the usage of the VH gene(s) identical or closely related to that of MOPC141 during early B-cell development.
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