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R Tobes

Publications and source records attributed to R Tobes.

4 recordsLinked to original sources
PubMed1997

Allele walking: a new and highly accurate approach to HLA-DRB1 typing. Application to DRB1*04 alleles.

We have developed a typing method, which can be used even in small laboratories, to produce a highly accurate and reliable allele assignment in any homozygous or heterozygous situation. We have called the method allele walking (AW) and it consists of sequential rounds of PCR-RFLP. After digestion, electrophoresis separates alleles positive for the mutation from the negative alleles; the cleaved fragment is then recovered from the gel and analyzed for mutations at another codon. In this way, AW is able to positively ascertain which mutations are in the same chromosome (cis-linkage) and assigns alleles independently from each other. Artificial sites are created in the PCR step in order to positively detect substitutions not naturally recognized by any of the existing or convenient enzymes. We report the application of AW for typing the 22 DRB1*04 alleles. The first PCR-RFLP round groups DRB1*04 alleles. Subsequently, the mutations at codons 86, 74, 71, 57 and 37 can be analyzed for the unambiguous assignment of the majority of the alleles. Additional polymorphisms at different codons can be assayed to resolve any undetermined alleles. The viability of all the restriction sites used as well as the feasibility of AW were successfully tested.

Alleles↗
PubMed1997

The tetramer model: a new view of class II MHC molecules in antigenic presentation to T cells.

Crystallographic studies suggest a plausible divalent interaction between T-cell receptor (TCR) and MHC class II molecules. In addition, biochemical data suggest that these divalent MHC molecules are preformed at the membrane of the antigen-presenting cell. The tetramer model is based on these preformed tetrameric class II molecules that can be loaded with identical or different peptides in their two grooves. This enables divalent class II molecules to deliver two different messages to T cell: 1) a two-peptide message, in which the tetramer with two identical peptides is able to cross-link two TCRs triggering full activation of a T cell. At the thymic level we propose that this message induces negative selection; or 2) a one-peptide message: only one of the peptides loaded in the class II tetramer is able to interact with that TCR. This message would be involved in triggering partial activation phenomena in mature lymphocytes, whereas in thymocytes this message would mediate positive selection. Since high concentrations of a peptide would favor the load of tetramers with identical peptides, the tetramer could therefore be viewed as a quantitative-qualitative transducer that would trigger different responses depending on the concentration of antigenic peptides.

Animals↗