Routine HLA-B genotyping with PCR-sequence-specific oligonucleotides (PCR-SSO) detects eight new alleles: B*0807, B*0809, B*1551, B*3529, B*3532, B*4025, B*5304 and B*5508.

This paper describes eight new alleles (B*0807, B*0809, B*1551, B*3529, B*3532, B*4025, B*5304 and B*5508) that have been found by routine HLA-B genotyping with sequence-specific oligonucleotides (SSOs). All of the new alleles have variations which cause changes in residues that occur within antigen binding pockets and T-cell recognition sites of the antigen. The new polymorphisms within these new alleles may affect the nature and specificity of peptide binding and cause differential T-cell activation, which may have an affect in transplantation.

A new HLA-A*02 allele, A*0234, detected by polymerase chain reaction using sequence-specific primers (PCR-SSP).

A novel HLA-A*02 allele, A*0234, was identified in a potential unrelated bone marrow donor typed by polymerase chain reaction using sequence-specific primers (PCR-SSP). Positive reactions obtained upon testing with PCR-SSP did not fit any known combination of alleles indicating the possible presence of a novel allele. Sequencing of clones from this individual revealed the presence of a novel allele, HLA-A*0234. The sequence of exons 2, 3 and part of exon 4 showed that A*0234 differed from A*02011 by a single nucleotide in exon 2 at position 282 (C to G). The nucleotide substitution results in an amino acid change at residue 70 (Histidine to Glutamine) in the alpha1 domain.

Confirmation of the detection of HLA-A*0104N in a family: a PCR-SSP reaction for the allelic detection of HLA-A*0104N.

The allele A*0104N has been detected in a family with a patient requiring a bone marrow transplant. The allele was found as a consequence of a discrepant result when family members were typed using serology and polymerase chain reaction using sequence-specific primers (PCR-SSP). Serological typing gave an apparent HLA-A 'blank' while PCR-SSP revealed the presence of an A*01 allele in three family members who were serologically negative for A1. Sequencing-based typing (SBT) was then used to establish that the allele was A*0104N. A PCR-SSP reaction was subsequently designed and used for the allelic detection of A*0104N. The study highlights the potential risks involved if molecular technology is used for typing, unless all non-expressed alleles are specifically detected.

Candidate regions for a testicular cancer susceptibility gene.

Epidemiological data suggest the presence of a susceptibility gene for testicular cancer in some families. Families with multiple cases of testicular cancer are rare and almost all those reported have only two affected members. We have performed a sib-pair analysis on 35 families in which there are either two or three affected brothers. These families have been typed for 220 autosomal microsatellite markers spaced 10-20 cM throughout the genome. Six regions which gave a LOD score of more than 1.0 on formal linkage analysis or a P value of 0.05 or less using a non-parametric approach are considered as candidate regions for a susceptibility gene. Of particular interest is one region on chromosome 4. Two neighbouring probes in this region both scored positively with LOD score of 2.60 on multipoint analysis. An International Testis Cancer Linkage Consortium has been formed to pool resources and will investigate these findings further with the world-wide collection of families.

HLA-A locus DNA typing of the 4AOH cell panel by arms PCR.

As part of the Fourth Asia-Oceania Histocompatibility (4AOH) Workshop, the authors have demonstrated a method of DNA-based tissue typing of the HLA-A locus using ARMS-designed primers in a panel of specific PCR reactions. The study was carried out blind under Workshop conditions and the results confirm the method as an accurate means of determining HLA-A locus tissue types.

Familial testicular cancer: a report of the UK family register, estimation of risk and an HLA class 1 sib-pair analysis.

Forty-two families with two or more cases of testicular cancer have been reported to the UK Register for Familial Testicular Cancer, comprising two pairs of identical twins, 27 sets of other brothers (25 pairs, two triples), nine father-son pairs, two pairs of first cousins and two uncle-nephew pairs. In total 91 testicular tumours are described in 86 individuals (42 (46%) pure seminoma, 49 (54%) other germ cell tumours). The median age at diagnosis in these patients was significantly younger than that in a comparable series of non-familial patients (29 c.f. 32.5 years, P less than 0.01). In a case-control comparison of 794 testicular cancer patients, eight patients (1.0%) had a brother and four patients (0.5%) had a father with a previous diagnosis of testicular cancer at the time of their own diagnosis (and these families are all included in this report). Two out of 794 controls (0.3%) had a first degree relative with testicular cancer. The cumulative risk to a brother of a patient for developing testicular cancer by the age of 50 years was estimated to be 2.2% (95% C.I. 0.6-3.8%) which results in a relative risk of 9.8 (95% C.I. 2.8-16.7) in comparison with the general population. HLA Class I typing of 21 affected sib-pairs demonstrated four (19%) sharing two haplotypes, 13 pairs (62%) sharing one and four pairs (19%) sharing none. This did not differ significantly from the expected proportions of 25%/50%/25%. It is unlikely, therefore, that there is a major gene associated with testicular cancer predisposition within or closely linked to the major histocompatibility gene complex on chromosome 6.

Microcytotoxicity assay using mouse L cells transfected with human MHC class II genes. A method and its application.

Modifications of the standard microcytotoxicity assay make it possible to use this technique to screen both alloantisera and monoclonal antibodies with mouse L cells transfected with Class II genes. It is necessary to maintain a protein-rich environment in order to prevent nonspecific complement lysis. Selection of the complement itself is also an important factor, the best results being achieved using a commercially available complement that had previously been absorbed with mouse cells and used at a dilution of 1/8. Using this modified method with transfectants of DW2 origin we could show that alloantisera against DRw15 recognize the DRB1*1501 gene product, whereas broad DR2 sera react only with the DRB5*0101 product. This technique can be applied successfully to study the fine specificity of polymorphic monoclonal antibodies, as shown by the reactivity of HU-30 which binds to the LDR2b transfectant and not to the LDR2a, indicating that the antibody recognizes an epitope present on the DRB1 chain and not the DRB5 chain of DR2 cell lines.