Linked Life Data

15350519

Source:http://linkedlifedata.com/resource/pubmed/id/15350519

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1-2
pubmed:dateCreated
2004-9-7
pubmed:abstractText
Topography of epitopes of monoclonal antibodies (MAbs), identified as the mutual competition of the MAbs, can be valuable indicators for the biological functions of MAbs. However, the determination of topographical epitopes is not performed before the functional screening of MAbs, because the requirement for purifying and labeling of MAbs makes the mapping experiment difficult, particularly in the early stage of MAb production. Here we describe a new label-free competitive enzyme-linked immunosorbent assay (LFC-ELISA) for the rapid grouping of MAbs based on the topography of their epitopes. In the LFC-ELISA, the immune complex formed by a competitor, MAb#2, and an antigen is challenged by an indicator, MAb#1 that had been captured on the ELISA plate through a secondary antibody. The MAb#2-antigen immune complex is trapped by MAb#1 only if MAb#1 reacts with an epitope different from that of MAb#2. The immune complex (MAb#2-antigen-MAb#1) is detected with an enzyme-labeled reagent specific to a tag on the antigen. Our experiments using different anti-CD30 MAbs and a CD30-Fc fusion protein as the antigen revealed that the LFC-ELISA performed well with MAbs of different isotypes (IgG1, IgG2a, and IgG2b), and in a practical range of MAb concentrations (0.3-10 microg/ml) and affinities (0.9-13 nM of Kd). We obtained pairwise competition data from all 26 anti-CD30 MAbs. We then utilized a cluster analysis and a bootstrap method to analyze the competition data for grouping of the MAbs. This objective and automated analysis identified eight distinct topographical epitopes on CD30. The reactivity of the anti-CD30 MAbs in immunoblot, and their inhibiting activity on CD30-CD30-ligand binding correlated with the topographical epitopes. The results show that the LFC-ELISA combined with cluster analysis is a useful new method for grouping MAbs based on their topographical epitopes and can be used in the early stage of MAb production. One useful application is to identify MAbs reacting with different epitopes from a large number of MAbs so that the most appropriate MAbs can be selected for therapeutic use.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0022-1759
pubmed:author
pubmed:issnType
Print
pubmed:volume
292
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
141-55
pubmed:dateRevised
2004-11-17
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
Rapid grouping of monoclonal antibodies based on their topographical epitopes by a label-free competitive immunoassay.
pubmed:affiliation
Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, 37 Convent Drive, Room 5106, Bethesda, MD 20892-4264, USA.
pubmed:publicationType
Journal Article