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Isolation of human anti-c-erbB-2 Fabs from a lymph node-derived phage display library.

Abstract：

:An immunoglobulin phage display library constructed from a tumour-associated pericolic lymph node was panned against the extracellular domain of the oncoprotein c-erbB-2. Sixteen independent clones were confirmed as positive binders based on ELISA analysis of soluble Fabs. Nucleotide sequencing demonstrated that the V(H) region of 12 clones belonged to four different V gene families, and the clones demonstrated varying degrees of somatic mutation compared with germ-line sequences. Fab fragments were examined for cross-reactivity by ELISA and shown to be negative against a panel of irrelevant self and non-self antigens, including bovine serum albumin (BSA), mouse immunoglobulin, tetanus toxoid, heregulin-PE40-FLAG and insulin. Reactivity of Fabs in vitro was verified by immunocytochemistry, which showed binding to the c-erbB-2 over-expressing breast cancer cell line SKBR3 but not to the low-expressing cell line MDA-MB-231. We conclude that a single lymph node library of moderate diversity (2 x 10(7) kappa light chain and gamma heavy chain clones), when derived from an individual whose colorectal tumour over-expressed c-erbB-2, can be successfully panned to isolate a number of unique Fabs specific for this antigen. The nature of the anti-c-erbB-2 Fabs recovered from this library suggests that they may have resulted from a humoral immune response in the individual, and that in vivo antibody responses to tumour-associated antigens may be exploited in vitro for the production of tumour-specific recombinant antibodies.

journal_name

Clin Exp Immunol

journal_title

Clinical and experimental immunology

authors

Clark MA,Hawkins NJ,Papaioannou A,Fiddes RJ,Ward RL

doi

10.1046/j.1365-2249.1997.4161316.x

subject

Has Abstract

pub_date

1997-07-01 00:00:00

pages

166-74

issue

1

eissn

0009-9104

issn

1365-2249

journal_volume

109

pub_type

杂志文章

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