Abstract
The monoclonal antibody DLT15, specific for thymocytes and peripheral T-cells of the teleost fish Dicentrarchus labrax: (sea bass), was used to purify immunoreactive cells from blood and gut-associated lymphoid tissue. The purification was performed by immune-magnetic sorting of leucocyte fractions enriched by Percoll density gradient centrifugation, and the purity of the isolated cells was estimated by cytofluorimetric analysis. Following a single step, the percentage of DLT15-purified cells was 88 +/- 10% for gut-associated lymphoid tissue and 79 +/- 18% for blood leucocytes. DLT15-purified cells from gut-associated lymphoid tissue were employed for RNA extraction and cDNA synthesis. In RT-PCR experiments using as primers degenerate oligonucleotides corresponding to the peptide sequence MYWY and VYFCA of the trout TcR beta chain, a 203 bp product was amplified. When sequenced, the cDNA was found to show 60% nucleotide identity to the tr out TcRV beta 3. By 3'-RACE the cDNA was elongated to obtain the TcR constant region, with high similarity to other fish TcR sequences. These results strongly suggest that cells recognised by DLT15 are putative T lymphocytes. (C) 2000 Academic Press.
Original language | English |
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Pages (from-to) | 329-341 |
Number of pages | 13 |
Journal | Fish & Shellfish Immunology |
Volume | 10 |
Publication status | Published - 2000 |
Keywords
- fish
- sea bass
- Dicentrarchus labrax
- monoclonal antibody
- T-cells
- T-cell receptor
- TROUT ONCORHYNCHUS-MYKISS
- THYMOCYTE MONOCLONAL-ANTIBODIES
- RECEPTOR-BETA-CHAIN
- CYPRINUS-CARPIO L
- RAINBOW-TROUT
- LYMPHOCYTE HETEROGENEITY
- ANTIGENIC DETERMINANTS
- CHANNEL CATFISH
- TELEOST FISH
- IMMUNOCYTOCHEMICAL DETECTION