Identification of monoclonal antibodies against 2,4-D herbicide by ELISA and DNA sequencing.
Seven hybridoma clones, E2/G2, E2/B5, E4/C2, G5/E10, F6/C10, B5/C3, and B7, produced within one fusion experiment in 1991 and the clone E4/C2 originated from 1995 were characterized by sequencing of genes coding for variable domains of the antibodies against 2,4-D herbicide. Amino acid sequences of selected antibodies, deduced from DNA analysis, were confirmed by mass spectrometry. Surprisingly, nucleotide sequence analysis of the clones E2/G2 and E2/B5, producing the most sensitive antibodies, proved to have sequence homology of their variable domains, although the IC(50) values determined for these antibodies 9 years prior to the DNA analysis were 2.0 and 8.2 ng/mL, respectively. The same findings arose from the comparison of the immunochemical to DNA data established for G5/E10, F6/C10, and B5/C3 clones producing antibodies with IC(50) values in the range of 26.3-43.1 ng /mL. The clone E4/C2, originating from the later fusion experiment, did not share nucleotide homology with any of the examined clones. Data obtained by ELISA, immunosensor, and DNA analysis within a 9 year period are discussed with respect to hybridoma stability, methodic artifacts, measurement reliability, and other possible factors influencing the result interpretation.
D015084:2,4-Dichlorophenoxyacetic Acid / Q000276:immunology*; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000737:chemistry / Q000235:genetics*; D000918:Antibody Specificity; D004247:DNA / Q000032:analysis; D004797:Enzyme-Linked Immunosorbent Assay; D006825:Hybridomas / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000737:chemistry; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000911:Antibodies, Monoclonal; D007135:Immunoglobulin Variable Region; D015084:2,4-Dichlorophenoxyacetic Acid; D004247:DNA