Analysis of amplified T cell receptor V beta transcripts by a non-isotopic immunoassay.
We have recently described a new colorimetric DNA enzyme immunoassay (DEIA) for detecting specific hybrids of complementary nucleic acids. This technology is based on an antibody that selectively recognizes double, but not single stranded DNA and therefore reveals the hybridization event independently from the DNA sequences. Most importantly, the test has an ELISA format and is very rapid and convenient for processing large numbers of samples. In the present report we have adapted this method to reveal the specificity of amplified T cell receptor V beta transcripts. V beta genes were amplified by polymerase chain reaction, using family specific primers and the specificity of the amplified products was determined by Southern blot and by DEIA. Our data demonstrate that DEIA had the same degree of sensitivity and specificity of conventional Southern hybridization. The possibility of analyzing amplified products with the simplicity of a conventional immunoassay should greatly facilitate the analysis of complex multigenic systems such as the T cell receptor and the immunoglobulin repertoire.
Consorzio per le Biotecnologie, School of Medicine, University of Brescia, Italy.
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Bettinardi
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Imberti
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Sottini
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Primi
D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis; D001345:Autoradiography; D001483:Base Sequence; D015139:Blotting, Southern; D017252:CD3 Complex; D004247:DNA / Q000096:biosynthesis; D003720:Densitometry; D005434:Flow Cytometry; D005803:Genes, Immunoglobulin; D006801:Humans; D007124:Immunoenzyme Techniques; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D007963:Leukocytes, Mononuclear / Q000276:immunology; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000235:genetics; D016133:Polymerase Chain Reaction; D012313:RNA / Q000032:analysis*; D011948:Receptors, Antigen, T-Cell / Q000032:analysis; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics*; D012680:Sensitivity and Specificity
D000945:Antigens, Differentiation, T-Lymphocyte; D017252:CD3 Complex; D007135:Immunoglobulin Variable Region; D015345:Oligonucleotide Probes; D011948:Receptors, Antigen, T-Cell; D016693:Receptors, Antigen, T-Cell, alpha-beta; D012313:RNA; D004247:DNA