bluuebunny/crossref_metadata_2025 huggingface dataset:
A family study of the association between insulin dependent diabetes mellitus, autoantibodies and the HLA system
Authors: Winearls B. C. , Bodmer J. G. , Bodmer W. F. , Bottazzo G. F. , McNally J. , Mann J. I. , Thorogood M. , Smith M. A. , Baum J. D.
Published: December 2008

A family study of the patients attending a puediatric diabetic clinic was undertaken. Seventy‐three percent of the index patients and their families provided samples for HLA typing and autoantibody analysis. The HLA types were analysed and correlated with the auto‐antibody data. Ways of predicting which siblings are at risk of developing insulin dependent diabetes mellitus (IDDM) were sought.

The antigen DR4 carries a higher relative risk and δ value (viz. 27 and 0.93) than any other antigen.

The data did not support the notion of heterogeneity of IDDM on the basis of HLA types or autoantibody status. However, the majority of patients with a younger age of onset were DR4/DR4 homozygotes.

The presence of ICA was a good marker for siblings at risk of developing IDDM. Some siblings who developed IDDM had ICA and CF‐ICA for long periods of time before developing diabetes.

The data supported an intermediate model of inheritance for IDDM.
Inhibition of cytotoxicity for screening a monoclonal antibody to HLA antigen
Authors: Hiroishi Shingo , Matsuyama Shigeyuki , Kaneko Toshiaki , Nishimura Yukiyo , Arita Junichiro
Published: December 2008

Cytotoxicity inhibition assay was established for the screening of a monoclonal antibody to HLA antigen. The assay involved the inhibition of typing cells with hybridoma culture supernatant and with F(ab')2 fragment of sheep anti‐mouse IgG. Using the assay and the conventional microcytotoxicity test, an anti‐HLA‐A2 monoclonal antibody was screened.
A high frequency of the A30, B18, DR3, DRw52, DQw2 extended haplotype in Sardinian celiac disease patients: Further evidence that disease susceptibility is conferred by DQ A1*0501, B1*0201
Authors: Congia Mauro , Frau Fulvia , Lampis Rosanna , Frau Rita , Mele Roberto , Cucca Francesco , Muntoni Francesco , Porcu Susanna , Boi Francesca , Contu Licinio , Nasa Giorgio La , Mulargia Marina , Pirastu Mario , Cao Antonio , De Virgillis Stefano
Published: December 2008

Abstract: This study characterizes by serological and molecular methods the HLA class I and class II alleles in a group of celiac disease children, their parents and a control group of Sardinian descent. We found the DR3‐DQw2 haplotype in all patients which was, in almost all cases (84%), associated with the HLA‐A30, B18, DR3, DRw52, DQw2 extended haplotype named “Sardinian haplotype” because of its frequency (12–15%) in this Caucasian population. This is the first time that this DQw2‐linked haplotype has been reported with such a high frequency in CD. However, no different distribution of “Sardinian haplotype” was found comparing CD patients with 91 haplotyped DQw2‐positive controls. This finding indicates that the DQw2 antigen in Sardinians is almost always associated with the A30, B18, DR3, DRw52, DQw2 extended haplotype. The DQA1 and DQB1 second exon sequence analysis of the B18,DR3 and B8,DR3 haplotypes showed the DQA1*0501 and DQB1*0201 alleles which shared the already published sequences. DPB1 subtyping showed the DPB1*0301 allele more frequently (p < 0.005) in CD patients but this difference was no longer significant when patients and controls, both heterozygous for the DR3‐DQw2 haplotype, were compared. We suggest that the divergent HLA extended haplotypes and DP allele associated with CD, described in different Caucasian populations, can be explained by the particular DQw2 linkage disequilibrium in each population.
HLA‐DRw52‐associated DRB1 alleles: Identification using polymerase chain reaction‐amplified DNA, sequence‐specific oligonucleotide probes, and a chemiluminescent detection system
Authors: Shaffer Arthur L. , Falk‐Wade Judith A. , Tortorelli Valeria , Cigan Amy , Carter Carolina , Hassan Kalpana , Hurley Carolyn Katovich
Published: December 2008

Abstract: Polymerase chain reaction (PCR) primers were designed to specifically amplify exon 2 of the DRw52‐associated DRB1 alleles for subsequent typing by sequence‐specific oligonucleotide probe (SSOP) hybridization and chemiluminescent detection. The DRw52 DRB1 group, encoding 22 of the 44 W.H.O. designated DRB1 allelic products, was divided by differential PCR with two polymorphism‐directed forward primers. Based on a polymorphism at codon 13, these forward primers separate the DRw52‐associated alleles into subsets; one comprised of the alleles of DR3/DRw11/DRw6 and the other of DRw8/DRw12/DRB1*1404. The DRB1 alleles in the latter subset were then defined by SSOP hybridization to the amplified DNA. The preferential amplification also resulted in SSOP definition of 15 alleles in the DR3/DRw11/DRw6 subset but some DRw11/DRw13 heterozygous allelic combinations were still unresolved. Two reverse PCR primers specific for the polymorphism at codon 86 were used to obtain amplified material to which SSOP reactivity provided definitive identification of the ambiguous heterozygotes.
Comparison of HLA class II alleles in Gypsy and Czech populations by DNA typing with oligonucleotide probes
Authors: Cerná Marie , Fernandez‐Viña Marcelo , Ivásková Eva , Stastny Peter
Published: December 2008

Abstract: Czechs (CZ) and Gypsies (GY) have lived in the same country for several centuries but seem to have different migratory origins. We have studied 99 CZ and compared them with 34 GY from an isolated group living in Czechoslovakia. DRBI, DRB3, DRB4, DRB5, DQAI, DQB1 and DPB3 alleles were determined by PCR followed by oligonucleotide hybridization. DRB 1*0701 was the most frequent DR allele in CZ (af=36.3%). In GY, after DRB1*0701, DRB1*1404 was the second most frequent allele (af=29.4%), while in CZ this allele was not found. Both groups differed also in the distribution of DR4 subtypes. In CZ, DRB 1*0401 was the most frequent subtype (50.0%) of DR4. In GY, DRB 1*0408 was the predominant type (44.4%). A new DQB1 pattern was found in GY, which we have called DQB1*3.4. This allele was similar to DQB 1*0301 in codon 26, but in codon 57 it resembled DQB 1*0302. All cells having this new allele were also positive for DRB1*0408. Four‐locus associations in CZ showed haplotypes similar to those previously found in other Caucasoid populations. In GY we observed more unusual haplotypes, some previously seen in the Orient, possibly reflecting GY ancestry.
Idiopathic dilated cardiomyopathy: lack of association between circulating organ‐specific cardiac antibodies and HLA‐DR antigens
Authors: Caforio A. L. P. , Martinetti M. , Schwarz G. , Bonifacio E. , Gavazzi A. , Graziano G. , Lorini R. , Cuccia M. , McKenna W. J. , Bottazzo G. F.
Published: December 2008

Abstract: Organ‐specific cardiac antibodies are serological markers of autoimmunity in dilated cardiomyopathy (DCM). HLA‐DR4 and possibly DR5 are immunogenetic markers of susceptibility in DCM, but it is not known whether they are associated with autoantibody production. We studied the frequency of HLA‐DR antigens and the presence of organ‐specific cardiac antibodies in 80 DCM Caucasian patients from Northern Italy. HLA‐DR typing was performed by serology; 289 healthy blood donors from the same region were tested as controls. HLA‐DR frequencies in DCM were also compared with VIII International Workshop control data for Italy. Cardiac antibodies were detected by indirect immunofluorescence on human heart. Skeletal muscle was used to identify cross‐reacting antibodies. The prevalence of cardiac antibodies in DCM was: organ‐specific 34% and skeletal muscle cross‐reactive 30%. The previously reported positive association between DCM and HLA‐DR4 was confirmed using either the controls from the same region (21.25% vs 10.73% p = 0.02, relative risk = 2.30) or from all of Italy (21.25% vs 12.3%, p = O.O3). HLA‐DR5 frequency was slightly but not significantly higher in DCM than in controls from the same region (46.25% vs 31.49% p = 0.02, relative risk of 1.87, p corrected = NS) or from all of Italy (46.25% vs 35.8% p = NS). HLA‐DR3 frequency was lower in DCM than in controls from the same region (12.50% vs 29.41% p = 0.003, relative risk of 0.36, p corrected = 0.03). This negative association was not confirmed using the control data from the whole of Italy (12.50% vs 16.5% p = NS). No significant association was found between HLA‐DR4, DR5 and/or DR3 and cardiac antibody status, age, sex, symptom duration. The lack of association between HLA‐DR and the presence of cardiac antibodies may be due to disease heterogeneity and/or reduction of antibody levels with disease progression, which are well‐recognised features of organ‐specific autoimmune conditions.
Population and family studies of HLA‐DR4 by use of oligonucleotide typing
Authors: Reed Elaine , Lupu Floria , McManus Peter , Seigle Robert , Suciu‐Foca Nicole
Published: December 2008

Abstract: Using PCR and DR4 group‐specific primers and SSO's we have examined DRB1*04 nucleotide polymorphism in a population of 123 DR4‐positive individuals (86 NAC, 27 Hispanics and 10 African Americans) from New York carrying a total of 134 DR4 haplotypes. We found that the distribution of DRB1*04 alleles on DR4 haplotypes differs in these three ethnic groups. In this relatively small population, certain alleles such as DRB 1*0406 and 0411 were encountered only in Hispanics, while others such as DRB 1*0403, 0408 and 0409 were found only in NAC (North American Caucasians). Such differences may be important in studies of HLA‐DR4 and disease associations. Evidence from MLC and PLT studies of an HLA‐BIDR crossover family, which was informative for the segregation of HLA‐DRB1*0406 and DRB1*0407, supports the concept that subtypes of HLA‐DR4 and/or associated HLA‐DP alleles elicit T‐cell alloreactivity, and may thus play a role in transplantation.
Canine leukocyte cell adhesion molecules (LeuCAMs): Characterization of the CD11/CD18 family
Authors: Danilenko D. M. , Moore P. F. , Rossitto P. V.
Published: December 2008

Abstract:The canine LeuCAM (CD11/CD18) family is characterized using a panel of newly developed anti‐canine LeuCAM monoclonal antibodies (mAb) as well as a cross‐reactive anti‐human CD11b mAb. The new anti‐canine LeuCAM mAb were developed by immunizing mice with purified canine CD11/CD18 antigen derived from an anti‐canine CD18 affinity column. Six mAb with reactivity to the canine LeuCAM family were cloned and characterized. These 6 included a new anti‐canine CD18 mAb, 2 anti‐canine CD11a mAb, 2 anti‐canine CD11c mAb, and a mAb which recognizes a novel canine CD11/CD18‐related macrophage antigen, designated LeuCAMmac, which is described in a separate report. The cell and tissue distribution of canine LeuCAMs was found to be similar to that reported in other species with a few notable exceptions. One prominent exception was the finding that canine CD11c, in contrast to human CD11c, was much more widely distributed on dendritic cells than it was on tissue macrophages. The molecular organization of the canine LeuCAM family was also found to be similar to that reported in other species, with the possible exception of the canine CD11b α chain, which may exist as several different species. It is anticipated that these anticanine LeuCAM mAb will prove useful in immunophenotypic studies of canine hemolymphatic system neoplasia.
Two new HLA DRB1 alleles found in African Americans: Implications for balancing selection at positions 57 and 86
Authors: Apple Raymond J. , Erlich Henry A.
Published: December 2008

Abstract: Two new DRB1 alleles have been identified (DRB1*0303 and DRB1*0805) in African Americans that differ from known DRB1 alleles only by a glycine to valine exchange at position 86. The novel DRB1*0303 allele, found in one individual, has the same DQA1*0401‐DQB1*04 haplotype as DRB1*0302, suggesting that it may be a recent diversification of *0302. The novel DRB1*0805 allele, identified in 4 individuals, was found on two haplotypes, sharing a DQA1*0501‐DQB1*0301 haplotype with DRB1*0804, and a DQA1*0102‐DQB1*0602 haplotype found with DRB1*0801 in some African populations. DRB1*0805 differs from *0804 only at position 57 and differs from *0801 only at position 86. Assuming that DRB1*0801 and DRB1*0802 are ancestral, based on their distribution in various human populations, DRB1*0805 may have been generated twice by two independent mutations or gene conversion events at each of these positions. Alternatively, DRB1*0805 may have arisen from a single gene conversion event (or mutation) and recombined to generate multiple DR‐DQ haplotypes. These findings increase the number of DRB1 allelic pairs that differ only at position 86 to 9, suggesting strong balancing selection at this position. A number of DRB1 alleles for DR8 and DR4 also differ only at position 57, a site previously postulated to be strongly influenced by balancing selection in DQB1 alleles by phylogenetic analysis.
Distribution of HLA antigens in Spanish Gypsies: A comparative study
Authors: de Pablo Rosario , Vilches Carlos , Moreno Maria E. , Rementería M. Carmen , Solís Rosario , Kreisler Miguel
Published: December 2008

We have studied the HLA‐class I and class II antigen distribution in a sample of 75 Spanish Gypsies and 74 Spanish non‐Gypsies by serology, restriction fragment length polymorphism, and protein chain reaction and hybridization with allele‐specific oligonucleotide probes. When both population samples are compared, we find that Gypsies have a statistically significantly higher frequency of A1, A11, B61, Cw6, DQ5 and haplotypes DR16 DQ5 Dw21 and DR14 DQ5 Dw9 DR52b. Frequency of A3, A29, B44, DR4, DQ2, DQ8 and haplotypes DR1 DQ5 and DR7 DQ2 DB17 DR53 are significantly lower in this ethnic group. The analysis of the serological data in the two populations demonstrates that Cw6 can be split into long Cw6 (Cw6.1) and short Cw6 (Cw6.2). Haplo‐type A1‐Cw6‐B61‐DR14‐DQ5 is the most characteristic in Gypsies, with a frequency of 13%. Estimation of the genetic distances shows that Spanish Gypsies are closer to Indian Caucasoid populations than to the Spanish non‐Gypsy population. HLA data support the proposed historical origin of this ethnic group.
Polymorphism of HLA in the Romanian population
Authors: Reed Elaine , Ho Eric , Lupu Floria , McManus Peter , Vasilescu Rodica , Foca‐Rodi Aurica , Such‐Foca Nicole
Published: December 2008

Abstract: We have investigated the HLA‐class I and class II polymorphism in a population of 83 Romanians using conventional serology together with PCR amplification and oligonucleotide typing of HLA‐class II genes. Romanians show a higher frequency of HLA‐A11, B13, B18, B37, B39, B51 and DR2 than other European populations. HLA‐DRB1*1501 and 1601 account for the high frequency of the serologic specificity DR2. In Romanians, HLA‐DR2 is in linkage disequilibrium with HLA‐B18 and HLA‐Bw52 rather than with HLA‐B7 as is the case in other Europeans. Unexpected HLA‐DR2 haplotypes include HLA‐DRB1*1502, DQA1*0102, DQB1*0601; HLA‐DRB1*1602, DQA1*0102, DQB1*0502. Other unusual haplotypes include HLA‐DRB1*0405, DQA1*03, DQB1*0302; HLA‐DRB1*1305, DQA1*0103, DQB1*0603; and HLA‐DRB1*1405, DQA1*0101, DQB1*05032. Analysis of the genetic distance between Romanians and other Europeans who have been studied serologically are consistent with the hypothesis that Romanians descend from Roman ancestors who colonized Dacia between the 1st century B.C. and 1st century A.D.
Oligotyping of Italian celiac patients with the 11th International Histocompatibility Workshop reagents*
Authors: Ferrante P. , Petronzelli F. , Mariami P. , Bonamico M. , Mazzill M. C.
Published: December 2008

Abstract: Oligotyping for HLA‐DRB1, DQA1 and DQB1 specificities has been performed on PCR‐amplified DKA from 55 Italian celiac children, living in Rome. and 50 blood donors. 52.6% of CD patients were DR3;DQ A1 *0501; DQB 1 *0201 ‐positive versus 14% of controls (RR = 6.85) and 34.5% were DR5,7;DQA1*0501;DQB1*0201‐positive versus 2% of controls (RR = 25.86). 7 patients (12.7%) were negative for the DQA1 *0501/B1 *0201 dimer: 3 of them were DR4 (5.4%) and the others typed as DR1,5; 1,7: 5,7 and w6,7. No patient was negative for both DQA1*0501 and DQB1*0201 alleles.
Alloantisera can detect some of the variants of HLA DRw13 identified by oligotyping*
Authors: Gebuhrer L. , Lambert J. , Labonne M. P. , Mallet I. , Freidal A. C. , Betuel H.
Published: December 2008

Abstract: Oligotyping has revealed a considerable polymorphism, pf HLA DRwl3; so far, 5 alleles coded by DRBI have been identified. An even greater number of haplotypes are apparent when considering the association of DRw13 with alleles coded by DRB3 and DQBI. Serology can now define some of these variants which had formerly escaped detection. We have tested by serology and by oligoprobes 66 genotyped Caucasoid cells comprising the known DR and DQ alleles. Among the 28 DRw6 cells, 10 different haplotype patterns were selected, of which 8 concern DRwl3 and 2 DRwl4. Four variants of DRw13 were represented: DRB1 *1301, * 1302. *1303 and 1305 associated with usual and with uncommon alleles belonging to DRB3 and to DQBI. We have centered our analysis on 19 sera of the XIth Workshop defining DRw6. Two sera (639. 826) were noteworthy since they reacted with certain subtypes of DRwl3 and with DRBI *1102, *1103 only, the other DR specificities remaining negative.
Rapid DNA typing of class II HLA antigens using the polymerase chain reaction and reverse dot blot hybridization
Authors: Buyse I. , Decorte R. , Baens M. , Cuppens H. , Semana G. , Emonds M.‐P. , Marynen P. , Cassiman J.‐J.
Published: December 2008

A nonisotopic oligotyping method using reverse dot blot hybridization was developed for HLA class II DQA1, DQB1, DPB1, DRB1, DRB3, DRB4, DRB5 alleles. The polymorphic second exon of the different genes was amplified by the polymerase chain reaction (PCR). For each gene the amplified DNA was hybridized at stringent conditions to membrane‐bound sequence‐specific oligonucleotides (SSOs) and visualization of positive signals was done by chemiluminescence. A combination of 11, 18, 23 and 31 SSOs was designed to identify 9/13 DQAI, 16/17 DQB1, 23/24 DPB1 and 50/55 DRB1, 4 DRB3, 1 DRB4, 3/4 DRB5 alleles respectively. For the DRB1 locus, an additional DRB1*04 groupspecific PCR was developed to make discrimination between the DR4 alleles possible in different heterozygous combinations. The procedure described here provides rapid and nonisotopic genotyping of heterozygous samples from a variety of sources and can be applied for tissue typing, disease susceptibility studies and forensic medicine.
Difference in HLA‐linked genetic background between mixed connective tissue disease and systemic lupus erythematosus
Authors: Dong Rui‐Ping , Kimura Akinari , Hashimoto Hiroshi , Akizuki Masashi , Nishimura Yasuharu , Sasazuki Takehiko
Published: December 2008

We have typed 64 Japanese patients with mixed connective tissue disease (MCTD) and 53 Japanese patients with systemic lupus erythematosus (SLE) for HLA‐DRB1, DRB3, DRB4, DRB5, DQA1, DQB1, and DPB1 genes by the HLA‐DNA typing method using the PCR‐SSOP technique. Frequencies of HLA‐DRB1*0401, DRB1*0901, DRB4*0101, and DQA1*03 were increased and those of HLA‐DRB1*0405 and DQB1*0401 were decreased in the patients with MCTD, while the frequencies of HLA‐DRB1*1501, DRB5*0101, and DQB1*0602 were increased in the patients with SLE. The typing results suggest that susceptibility to MCTD is strongly associated with the HLA‐DRB1*0401‐DRB4*0101‐DQA1*03‐DQB1*0301 haplotype, and that to SLE is associated with the HLA‐DRB1*1501‐DRB5*0101‐DQA1*0102‐DQB1*0602 haplotype. The observation that the MCTD‐associated HLA alleles are distinct from the SLE‐associated ones may support the clinical entity of MCTD different from SLE.
The monoclonal antibody TAL16.1 recognizes the aspartic acid residue at position 70 in DRB gene products
Authors: Sadler A. M. , Heyes J. M. , Marsh S. G. E. , Krausa P. , Reynolds G. E. , Bodmer J. G.
Published: December 2008

A polymorphic monoclonal antibody (TAL16.1), raised against a mouse L‐cell transfectant expressing the human DRB5*0101 gene from the HLA‐DR15(2) Dw2 DR51 haplotype was shown to have a complex pattern of reactivity to DRB gene products. The antibody bound to a transfectant expressing the DRB5*0101 allele against which it was produced but not to a transfectant expressing the DRB1*1501 allele. These alleles of the DRB1 and DRB5 genes are usually coexpressed on DR15(2) Dw2 DR51 cells. A comparison of the HLA‐DRB amino acid sequences of reactive and non‐reactive cells identified an aspartic acid residue at position 70, conserved in all antibody‐positive cells and absent in antibody‐negative cells, which was postulated as being responsible for conferring the specificity of the antibody. The aspartic acid residue at position 70 is present in DRB5*0101 and DRB5*0102 alleles but absent in DRB5*0201 and DRB5*0202 alleles, allowing the antibody to distinguish between these splits of the DR51 serological specificity. TAL16.1 also binds to the product of the DRB1*0103 allele and discriminates between cells with a DR103 specificity and the other DR1 subtypes, DRB1*0101 and DRB1*0102. In this report the value of transfectants as immunogens for use in the production of monoclonal antibodies of predetermined specificity and as tools for the fine mapping of antibody specificity is discussed.
Major Histocompatibility Complex class II DQ diversity in Rhesus macaques
Authors: Slierendregt Bastiaan L. , Otting Nel , Jonker Margreet , Bontrop Ronald E.
Published: December 2008

By the use of restriction fragment length polymorphism analysis 10 Taq I fragments could be identified for the MhcMamu‐DQA1 region. A strong correlation exists between the occurrence of Mamu‐DQA1/Taq I fragments and Mamu‐DAQ1 allelic sequence variation. Most restriction fragments correspond with a unique Mamu‐DQA1 allele, with one exception being the Taq I 4.5 kb fragment that is associated with two Mamu‐DQA1 alleles. The RFLP technique allowed the identification of 15 Mamu‐DQB1/Taq I restriction fragments, whereas sequence analysis has permitted the characterization of at least 20 different Mamu‐DQB1 alleles. In this communication two unpublished Mamu‐DQB1 sequences are described. For Mamu‐DQB1, on only four occasions was it possible to demonstrate a correlation between a certain fragment and an allelic sequence. These analyses, performed on material from truly homozygous animals, allowed us to define which combinations of Mamu‐DQA1 and ‐DQB1 molecules form heterodimers at the cell surface. In addition, these studies are helpful in typing non‐human primate species that are used in biomedical research.
Rapid DNA typing for Class II HLA antigens: Subtyping of DRw52‐associated DRB1 alleles
Authors: Horne C. , Keown P. A.
Published: December 2008

Serologic typing for MHC class II antigens is incapable of identifying important subtypes for certain DRB1 alleles and occasionally leads to errors of assignment, particularly with the DR antigens associated with DRw52. To simplify DNA typing of DRw52‐associated DRB1 alleles, we have developed a new rapid method using PCR‐RFLP. The PCR‐RFLP method is based on allele‐specific amplification followed by digestion of PCR‐amplified DNA with restriction enzymes. Group‐specific amplification of the second exon of DR3, DR5, DR6 and DR8 was achieved using a 5’primer specific for the first hypervariable region sequence common to all alleles in this group and generic 3’primers. Human genomic DNA was amplified in a Perkin‐Elmer Thermocycler. The presence of a 265 bp fragment was confirmed by agarose gel electrophoresis. Restriction enzyme digestion using Rsa I followed by polyacrylamide gel electrophoresis gave a pattern unique for some alleles and placed the remainder in subgroups. Digestion of the PCR product with one or two of the following enzymes (Asp 700, Hae II, Mnl I, Mbo II, Ksp I and Hph I) permitted the identification of 21 of the 22 alleles. DRB1*1103 and DRB1*1104 are not distinguished by this method and can be distinguished by SSOP or by using a specific 3’primer. For some heterozygous combinations, additional primers are used to provide full subtyping. This method provides a rapid and less costly alternative to PCR‐SSOP for DRw52 subtyping in the smaller laboratory as only one amplification is required (two primers) for the majority of samples. The patterns produced by restriction digest are easy to interpret and complete subtyping using three additional primers can be accomplished in less than 2 days. This method should significantly improve the selection of unrelated donors for bone marrow or solid organ transplantation and facilitate immunogenetic studies in autoimmune diseases.
Typing of serum‐soluble HLA‐B27 antigen by ELISA
Authors: Pouletty Philippe , Chang Chin , Kalil Jorge , Atwood Elizabeth , Ferrone Soldane , Shimizu Bob , Howson William , Mazaheri Reza , Villano Bart Del , Grumet Carl
Published: December 2008

Abstract: An ELISA using serum as soluble HLA antigen source was developed for HLA‐B27 typing. Two sandwich assays were run in parallel. The first assay utilized a monoclonal antibody (mAb) reacting with a determinant expressed by both HLA‐B7 and B27 antigens; the other assay utilized a mAb reactive with HLA‐B7 antigens but not with HLA‐B27 antigens. After incubation with serum samples, bound HLA antigen was detected using an anti‐β2m antibody conjugated to peroxidase and a chromogenic substrate. Absorbance of each well was measured at 490 nm. Based on analysis of absorbances obtained with panels of specimens of known HLA phenotypes, a mathematical algorithm was developed to derive the specimen HLA‐B27 phenotype from its ELISA absorbance values. Despite the lack of monospecific mAb, an accurate HLA‐B27 typing was possible. 362 specimens (including 151 HLA‐B27‐positive) were tested. Agreement between microlymphocytotoxicity and ELISA was 99.2%. No correlation between the level of HLA‐B27 antigen reactivity and the amount of total HLA class I antigen in serum was observed. This report demonstrates the possibility of using serum‐soluble HLA antigen and ELISA technology for histocompatibility testing. The assay offers several significant advantages over microlymphocytotoxicity: no need for cell preparation, batch testing capabilities and objective, reproducible interpretation of results.
Gametic association of HSP70‐1 promoter region alleles and their inclusion in extended HLA haplotypes
Authors: Cascino I. , D'Alfonso S. , Cappello N. , Giordano M. , Pugliese A. , Awdeh Z. , Alper C. A. , Richiardi P. Momigliano
Published: December 2008

Abstract: Gametic associations of a three‐allele polymorphism of the HSP70‐1 promoter region were analyzed in a random North Italian population, in 69 HLA homozygous cell lines and in 29 families in Boston, all typed for HLA class I, class II and complement alleles. Significant phenotypic associations were detected in the random population between HSP70‐1 alleles and several HLA markers carried by extended haplotypes. The inclusion of HSP70‐1 alleles in extended haplotypes, suggested by population analysis, was confirmed in genotyped cells, including 10W HLA homozygous cell lines and families, selected for the presence of the whole set of alleles reported for conserved extended haplotypes. Every tested extended haplotype was exclusively associated with a given HSP70‐1 allele, except those carrying DR1. HSP70‐1 C was included in both [HLA‐B8, SC01, DR3] and [HLA‐B18, F1C30, DR3] extended haplotypes, accounting for the previously observed strong association with DR3. In addition the same allele was found on the [HLA‐B13, SC31, DR7], on the [HLA‐B62, SB42, DR4] and on the [HLA‐B60, SC02, DR13] extended haplotypes. The HSP70‐1 A allele was carried by all DR4+ extended haplotypes except the one above cited. HSP70‐1 B correlated with DR10, DQB1*0501 and BF*F. Thus the HSP70‐1 promoter alleles provide new precisely located markers of extended haplotypes.
Quantitation and characterization of plasma HLA in neonates of different gestational ages
Authors: Puri Suksham , Shieh D. C. , Canavan Allyson , Kao K. J.
Published: December 2008

Abstract: The functional significance of plasma HLA class I antigens is unclear. They are thought to have an immunomodulatory role and be tolerogenic in transplant settings including the materno‐fetal semi‐allograft. There is, however, no available data on the concentrations of soluble HLA class I antigens in fetuses or newborns. We therefore determined plasma HLA class I antigen levels in 93 neonates born at different gestational ages and compared them to those in 66 healthy adults. The mean plasma HLA concentration in cord blood obtained from these neonates (0.30 ± 0.15 μg/ml, mean ± SD) was significantly lower (p<0.0001) than in the adults (0.77 ± 0.44 μg/ml). No correlation between the plasma HLA levels and the gestational ages of the neonates was detected. Characterizing the plasma HLA class I antigens by immunoprecipitation and immunoblotting, four different molecular weight forms, 44, 39, 36 and 34 kDa, were recognized. Their distribution in neonates was not different from that in adults. Since the circulating leukocytes are a probable source of plasma HLA class I antigens, we measured the surface HLA expression on leukocytes in 4 neonates and 4 adults by immunofluorescent flow cytometry. The fluorescence intensities on neonatal granylocytes and lymphocytes were 50% of those on corresponding adult cells. This finding suggests that the reduced HLA expression by neonatal leukocytes may be partially responsible for the lower concentration of HLA class I antigens in neonatal plasma.
A monoclonal antibody with specificity to the HLA‐DR1 and ‐DR51 antigens
Authors: Loh M. T. , Chan S. H. , Ren E. C.
Published: December 2008

Abstract: A monoclonal antibody 137BL7 raised against purified DR1 protein was shown to bind specifically to 5/5 DR1, 18/18 DR2 cells and 0/23 non‐DR1,2 cells by cell‐EIA. Further analysis by FACS using HLA‐transfectants revealed that 137BL7 also bound specifically to the DRB5 transfectants in addition to the expected DR1 transfectants. However, it did not bind to the DR2 (DR15) transfectants, showing that cross‐reactivity with DR2 cells lies with the DRB5 (DR51) rather than the DRB1 gene product. A comparison of the HLA‐DR amino acid sequences of DR1 and DR51 antigens revealed a common glutamic acid residue at position 96, which may form the putative binding epitope of this mAb.
The MLC assay as a test for HLA‐D region compatibility between patients and unrelated donors: Results of a national marrow donor program involving multiple centers
Authors: Mickelsen Eric M. , Bartsch Glenn E. , Mansen John A. , Dupont Bo
Published: December 2008

Abstract:Mixed lymphocyte culture (MLC) assays from 763 patients and prospective unrelated marrow transplant donors identified through the National Marrow Donor Program (NMDP) were analyzed for their overall utility in measuring HLA‐D region Compatibility. The assays were performed at 31 different transplant centers (range, 1 to 197; median, 9 assays per center). A total of 325/763 (42.6%) of the tests were judged to be uninterpretable. either due to lack of sufficient control cells included in the assay (89 tests) or to insufficient reactivity by patient and/or donor cells (236 tests). Among the 438 tests that could be interpreted, HLA‐Dw phenotyping with HLA‐D homozygous cells was performed for a subset of 190. The relative response (RR) values from these 190 tests, however, were not clearly separable into distinct populations; i.e., RR values corresponding to Dw identity versus nonidentity between patient and donor could not be reliably discriminated. The predictive value of a nonreactive MLC for Dw identity was calculated to be 0.91 for RRs of ≤ 20%, while the predictive value of a reactive MLC for Dw nonidentity was 0.35 for RRs of > 20%. These results, based on an analysis of data submitted from multiple transplant centers testing patients who had a variety of hematologic disorders, suggest that the MLC assay is a relatively imprecise method for determining HLA‐D region compatibility between patient and prospective unrelated marrow donor.
A novel HLA haplotype containing a DRB5 gene associated with the DRB1 *0103 allele
Authors: Perrier P. , Leotard B. , Henry M. A.
Published: December 2008

Abstract: A new hybrid haplotype, a DRB1 *0103 allele associated with a DRB5 *0101 allele, was found in a French Caucasoid family and has been described here. When these cells were typed by serology, contrarily to cells with the DR1 + 2s haplotype, they did not seem to be triplets. The reactivity of these cells with the DR2 allosera led to a False serological HLA‐DR typing. RFLP analysis and DNA oligotyping after DR1‐DRB1, DR1‐DRB1 and DRB5 group‐specific amplifications showed that there was no DR2‐DRB1 product in these cells and demonstrated the segregation of a DR103 DR51 haplotype in the family.
Genes within the HLA class II region confer both predisposition and resistance to primary biliary cirrhosis
Authors: Begovich Ann B. , Klitz William , Moonsamy Priscilla V. , Van de Water Judy , Peltz Gary , Gershwin M. Eric
Published: December 2008

Abstract: Nonradioactive sequence‐specific oligonucleotide probes for the polymorphic HLA class II genes have been used to type samples from 51 Caucasian patients with the autoimmune liver disease, primary biliary cirrhosis, and 240 Caucasian controls. Although the allelic distribution at the DPB1 locus showed no significant variation between patients and controls, there was heterogeneity in the distribution of DR‐DQ haplotypes where the frequency of the DRB1*0801‐DQA1*0401/0601‐DQB1*04 haplotype was significantly increased in the patients, suggesting it confers susceptibility to this disease. Two other haplotypes, DRB1*1501‐DQA1*0102‐DQB1*0602 and DRB1*1302‐DQA1*0102‐DQB1*0604, were significantly reduced in the patients, suggesting they confer protection. Tests of the individual loci show that resistance to this disease is most strongly associated with the DQA1*0102 allele shared by both protective haplotypes. Due to linkage disequilibrium it is unclear whether multiple genes or a single locus on the susceptible DR8 haplotype are needed for predisposition. These data show that distinct HLA class II alleles confer both predisposition and resistance to PBC and provide insight into the role that these genes may play in the immunopathogenesis of this disease.
Prevention of graft‐versus‐host disease in DLA‐haplotype mismatched dogs and hemopoietic engraftment of CD6‐depleted marrow with and without cG‐CSF treatment after transplantation
Authors: Schumm Michael , Günther Wolfgang , Kolb Hans‐Jochem , Rieber Peter , Büttner Matthias , Voss Christiane , Kremmer Elisabeth , Reitmeier Peter , Thierfelder Stefan , Wilmanns Wolfgang
Published: December 2008

Abstract: Prevention of graft‐versus‐host disease by depletion of CD6‐positive T cells was studied in the dog. Donors were DLA‐homozygous, recipients DLA‐heterozygous with one DLA haplotype identical to the donor. Seven control dogs received untreated marrow and died of GvHD after full hemopoietic recovery within 28 days of transplantation. For prevention of GvHD, immunomagnetic separation of T cells with a monoclonal antibody against human CD6 that crossreacted with canine T cells was evaluated. Depletion of CD6‐positive cells depleted CD4‐positive cells completely, but only part of CD8‐positive cells and DR‐positive cells. CD6‐depleted marrow exhibited strong nonspecific “natural” suppression of the generation of cytotoxic T cells in vitro. Eleven dogs received CD6‐depleted marrow. Only 1 dog developed GvHD and died. Sustained engraftment was seen in 8 dogs. Hemopoietic recovery was delayed and slower after transplantation of CD6‐depleted marrow than after transplantation of untreated marrow. Four of these dogs were treated with G‐CSF, and this accelerated the recovery of leukocytes, but did not prevent rejection. Chimerism was mixed in 7 of 10 evaluable dogs and 1 dog recovered its own hemopoiesis 2 years after transplantation. CD6 depletion prevents GvHD across a DLA‐haplotype difference, but rejection and mixed chimerism may occur. Treatment with G‐CSF accelerates leukocyte recovery, but cannot prevent rejection.
Generation and characterization of a human monoclonal IgG antibody specific for HLA‐B12
Authors: Siemoneit K. , Wölpl A. , Wegener S. , Ernst M. , Sonneborn H.‐H. , Goldmann S.F.
Published: December 2008

Abstract: We describe here the generation and characterization of a human monoclonal IgG antibody (UL/F14) specific for HLA‐B12. The antibody is suitable for complement‐dependent lysis on lymphoblastoid cell lines and stimulated peripheral blood mononuclear cells. UL/F14 competes for antigen binding with an HLA‐B12 human monoclonal IgM antibody and with a specific alloantiserum. Protein chemistry shows that the monoclonal antibody UL/F14 can precipitate solubilized HLA‐B12 molecules.
UN1, a murine monoclonal antibody recognizing a novel human thymic antigen
Authors: Tassone Pierfrancesco , Bond Heather , Bonelli Patrizia , Tuccillo Franca , Valerio Giuliana , Petrella Antonello , Lamberti Annalisa , Cecco Luigi , Turco Maria Caterina , Cerra Maria , Montagnani Stefania , Morrone Giovanni , Venuta Salvatore
Published: December 2008

Abstract: A murine monoclonal antibody (mAb) UN1 was produced on the basis of selective reactivity with human thymocytes. Characterization of UN1 by immunofluorescence gave a high intensity of labeling with the majority of human thymocytes. Expression was preferentially associated with immature thymocytes (CD3dim) compared to mature cells, whereas only a subpopulation of peripheral blood lymphocytes was weakly stained. No specific binding to monocytes or granulocytes was detected. The T‐cell lines HPB‐ALL, H9 and MOLT‐4 were all positively bound by UN1. Immunohistological staining of thymic tissues showed that mAb UN1 detected cells in both the cortex and medulla of fetal thymus, whereas the reaction in thymus samples from young children was mainly with medullar cells. By western blotting analysis, the antigen recognized by mAb UN1 corresponds to a membrane polypeptide with a molecular weight of approximately 120 kDa present on thymocytes and HPB‐ALL cells. The mAb UN1 was submitted to the 5th International Workshop and Conference on Human Leukocyte Differentiation Antigens, Boston, 1993. UN1 did not cluster in any of the old or new clusters of differentiation discussed at the conference, indicating its unique reactivity. Together with the data presented in this paper, this suggests that the UN1 antibody defines a previously undescribed molecule present on the cell surface of thymocytes and a minority of peripheral blood lymphocytes.
Role of the mixed lymphocyte culture (MLC) reaction in marrow donor selection: Matching for transplants from related haploidentical donors
Authors: Mickelson Eric M. , Guthrie Leigh Ann , Etzioni Ruth , Anasetti Claudio , Martin Paul J. , Hansen John A.
Published: December 2008

Abstract:The utility of the MLC assay as a test of HLA‐D region matching and predictor of acute graft‐versus‐host disease (GvHD) was evaluated in 157 patients receiving marrow grafts from HLA‐A, B identical related haploidentical donors. All donors and recipients were tested by HLA‐DR serology, by Dw phenotyping with homozygous typing cells (HTC) and by standard MLC. Ninety‐nine of the donor‐recipient pairs were mismatched for a serologically defined HLA‐DR antigen while 109 pairs were mismatched for the HLA‐D region by HTC typing. Donor anti‐recipient relative responses (RR) in MLC, corresponding to the GvHD vector in marrow transplantation, ranged from ‐4% to 100%, with a median of 25%. A comparison of reactivity in MLC with presence or absence of matching by Dw phenotyping, however, showed a significant overlap in the distribution of RRs from HLA‐Dw matched versus Dw mismatched pairs, suggesting that the MLC was not a reliable predictor of HLA‐Dw matching. Using an optimally‐defined cutoff of 3% RR, the MLC was correlated with risk of developing clinically significant grades II–IV acute GvHD (p = 0.03) but not with risk of developing severe grades III‐IV GvHD (p = 0.18). In contrast, matching by Dw phenotype was a significant predictor of GvHD, with Dw‐compatible transplant recipients less likely to develop either grades II‐IV (p = 0.004) or III‐IV (p = 0.036) GvHD than Dw‐incompatible transplant recipients. Overall, these results underscore the difficulty in using the MLC to measure HLA‐D region compatibility and predict the risk of severe graft‐versus‐host disease among patients receiving related haploidentical marrow grafts. HLA‐D (HTC) typing results correlate primarily with DRB compatibility, and with the advent of DRB1 allele matching by sequence‐specific oligonucleotide probes (SSOP) or by direct sequencing, the precision in donor matching achievable with these methods is far greater than with either HLA‐D typing or direct MLC testing.
A small test of a sequence‐based typing method: Definition of the B*1520 allele
Authors: Domena John D. , Little Ann‐Margaret , Arnett Kelly L. , Adams Erin J. , Marsh Steven G. E. , Parham Peter
Published: December 2008

Abstract:Santamaria et al. (Human Immunology 1993 37: 39–50) describe a method of sequence‐based typing (SBT) for HLA‐A, B and C alleles said to give “unambiguous typing of any sample, heterozygous or homozygous, without requiring additional typing information”. From SBT analysis, which involves determination of partial sequences of mixed alleles, these investigators reported that cell lines KT17 (HLA‐B35,62) and OLGA (HLA‐B62) from the reference panel of the 10th International Histocompatibility Workshop express novel variants of HLA‐B15 (B1501‐MN6) and HLA‐B35 (B3501‐MN7) respectively. To study further the novel alleles, we cloned and sequenced full‐length HLA‐B cDNA clones isolated from the KT17 and OLGA cell lines. We find that KT17 expresses B*3501, as assigned by SBT, and B*1501, the common allele encoding the B62 antigen. We were unable to confirm that KT17 expresses the novel B1501‐MN6 variant identified by SBT. For OLGA our analysis confirms the partial sequences obtained by SBT. Thus OLGA expresses B*1501 and a novel HLA‐B allele. The complete sequence of the latter shows it is a hybrid having exons 1 and 2 in common with B*1501 and other B15 subtypes and exons 3–7 in common with B*3501 and related molecules including B*5301 and B*5801. The novel allele has been designated B*1520 because of its sequence similarity with the B15 group; furthermore, serological analysis shows that the B*1520 product does not express epitopes in common with either B35, B53 or B58. The B*1520 heavy chain has a similar isoelectric point to A*3101; B*1520 was undetected by previous applications of isoelectric focusing because B*1520 and A31 are both expressed by OLGA. In conclusion, HLA‐B typing of two cell lines by cDNA cloning and sequencing gives concordant results with SBT for three of the four alleles. The cause of the discrepany for the fourth allele is unknown, however this finding indicates that the novel HLA‐A, B and C sequences emerging from SBT studies need independent verification.
A novel HLA‐A determinant recognized by a cytotoxic human hybridoma IgG1 monoclonal antibody (TrJ14)
Authors: Ge Jun , Bratlie Anne , Hannestad Kristian
Published: December 2008

Abstract: TrJ14 is a cytotoxic human IgG1Λ hybridoma mAb that recognized a novel HLA‐A epitope expressed by lymphoblastoid B cells that are homo‐ or heterozygous for A2, A3, A11, A30, A31, A33, A68 and A69. Based on these results, the HLA type of cell line TEM (10w9057) was retyped as A66. When peripheral blood T cells isolated freshly from 265 HLA‐typed normal individuals served as targets, TrJ14 killed cells expressing two TrJ14‐positive HLA‐A alleles, as well as the majority of cells having one TrJ14‐positive and one TrJ14‐negative HLA‐A antigen. However, TrJ14 failed to recognize or reacted weakly with most HLA‐A2 and ‐A3 heterozygous normal T cells when A2 or A3 was coexpressed together with a TrJ14‐negative antigen. The serological reactivity of TrJ14 correlated with the amino acid valine and aspartic acid at positions 76 and 77 of the αl‐domain helix. These amino acids were shared exclusively by all the identified TrJ14+ alleles.
HLA‐B16 antigens: sequence of the ST‐16 antigen, further definition of two B38 subtypes and evidence for convergent evolution of B*3902
Authors: Adams E. J. , Martinez‐Naves E. , Arnett K. L. , Little A.‐M. , Tyan D. B. , Parham P.
Published: December 2008

Abstract:The ST‐16 antigenic specificity of the HLA‐B locus is defined as a B39 variant of Mexican‐Americans. Nucleotide sequencing of cDNA shows the ST‐16 allele (B*3905) differs from B*39011 by a single substitution that substitutes tyrosine for aspartic acid at position 74 of the mature class I heavy chain. The complete coding region sequence for the common caucasoid allele encoding the B38 antigen has been determined. This B*3801 allele differs from B*3802 at two nucleotide substitutions within the Bw4 sequence motif. B*3801 and B*3802 may have been derived independently from B*39011 by conversion events with B alleles donating distinctive Bw4 motifs. A novel allele B*39022 derived from a Colombian Indian differs from the B*39021 allele of Japanese origin at two widely separated silent substitutions. Comparison of sequences for the known B16 alleles suggest that B*39021 and B*39022 were independently derived by recombination from B*39013 and B*39011 respectively.
Serology versus PCR‐SSP in typing for HLA‐DR and HLA‐DQ: a practical evaluation
Authors: Otten H. G. , Tilanus M. G. J. , Barnstijn M. , van Heugten J. G. , de Gast G. C.
Published: December 2008

Abstract: In this study, serological HLA‐DR and ‐DQ typing results were compared to typing results obtained with sequence‐specific primers in the polymerase chain reaction (PCR‐SSP). HLA‐DR typing was performed on a random Caucasian population consisting of 31 patients and 73 healthy individuals. Considering HLA‐DR1‐10, differences in typing results were found in 3 out of 73 healthy individuals and 8 out of 31 patients. When HLA‐DR1‐16 alleles were taken into account, differences in typing results were found in 11 out of 31 patients and 14 out of 73 healthy individuals. Typing results of PCR‐SSP, different from that of serology, were all confirmed by sequencing‐based typing of HLA‐DRB1 alleles. HLA‐DQ 1–3 typings were performed on 40 individuals consisting of 17 patients and 23 healthy individuals. Differences in typing results were found in 5 out of 17 patients and 1 out of 23 healthy individuals. From the results of this study it can be concluded that serology is a reliable technique, when restricted to identification of HLA‐DR1‐10 and HLA‐DQ1‐3 antigens in healthy individuals. By PCR‐SSP, however, reliable HLA‐DR1‐16 and ‐DQ1‐3 typings can be obtained both in patients and healthy individuals.
Distribution of HLA and haplotypes of Colombian and Jamaican black populations
Authors: Blank M. , Blank A. , King S. , Yashiki S. , Kuwayama M. , Fujiyama C. , Gongora D. , Zaninovic V. , Cranston B. , Hanchard B. , Imanishi T. , Manns A. , Blattner W. A. , Tajrma K. , Hayami M. , Fujiyoshi T. , Sonoda S.
Published: December 2008

Abstract: To investigate the genetic background of the black populations of Colombia and Jamaica, we determined HLA types of 78 Colombian and 98 Jamaican blacks from 2 different socioeconomic groups (Jamaican #1 and Jamaican #2) and estimated the frequencies of HLA genes and haplotypes. A phylogenetic tree based on the HLA gene frequencies revealed that Jamaican #1 and Jamaican #2 were distinct from each other, Jamaican #1 being closely related to the Colombian blacks and the Jamaican #2 being closely related to Senegalese and Zairean populations. Three‐locus HLA haplotypes of Colombian and Jamaican #1 blacks were an admixture between Africans and Caucasians or South American Indians, while Jamaican #2 blacks were relatively homogeneous and appeared to conserve African lineages. The major five‐locus HLA haplotypes were not shared among Colombian, Jamaican #1 and Jamaican #2 blacks. These results indicated that the black populations of Colombia and Jamaica were originated from African blacks and admixed variably with Caucasians and South American Indians to make genetic subpopulations in Colombia and Jamaica.