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PubMed1995

Evaluation of HLA antibodies with the PRA-STAT test. An ELISA test using soluble HLA class I molecules.

HLA-specific antibody, present before or after transplantation, may adversely effect graft outcome. Antibody testing by cytotoxicity (CYT) is laborious, requires viable lymphocytes, does not differentiate non-HLA cytotoxic antibody, and cannot be used readily on specimens from patients being treated with cytotoxic antibodies. We have evaluated PRA-STAT, an antibody screening kit that uses an ELISA test with soluble HLA class I molecules as targets. We performed 219 tests on a variety of serum specimens, 128 of which were also tested by CYT. There was a highly significant correlation (r = 0.78, P < 0.001) between PRA-STAT (PS) and CYT for the detection of IgG antibodies. Of 66 sera reactive in both assays, 18% had identical specificities defined in both, 27% were more reactive in PS than in CYT, 8% were more reactive in CYT, and 47% had different specificities in the 2 assays, with overlap in slightly more than half the cases. Of 13 sera reactive only in PS, 2 were from non-transfused, nontransplanted males with no evidence of lymphocyte-reactive antibody by antiglobulin tests. PS uses an IgG-specific conjugate, therefore IgM class I-specific antibodies cannot be identified--however, their presence does affect test outcome. This, as well as the panel composition and interlot reproducibility, are areas we believe need to be addressed. The PRA-STAT system is rapid, does not require viable cells or complement, and can be automated in part. Resolution of the problems identified here and availability of an IgM-specific conjugate should make this test system a valuable tool in histocompatibility testing.

Antibodies↗
PubMed1980

Recognition of a positive MLC reaction after four hours.

EMT indicator cells were incubated for 1 h in the supernatant of a 3 h MLC. Their electrophoretic mobility was then measured by an analytical, carrier-free electrophoresis system. The alteration of their mobility against a medium-standard was calculated in per cent and correlated with the conventional measured MLR-cpm. According to our results the correlation of the two quantities is statistically highly significant. This method could enable recognition of a positive or negative MLC after only 4 h. This fact could be of great importance for histocompatibility testing and transplantation.

Cell Movement↗
PubMed1978

Vertebral ankylosing hyperostosis (Forestier's disease) and HLA antigens in Pima Indians.

Vertebral ankylosing hyperostosis is a common skeletal disorder among Pima Indians. Prevalence rates of approximately 50% have been reported in Pima Indian males aged 44 or older. The HLA-B27 antigen is also found more commonly in Pima Indians than in Caucasians (18% versus 6%). Because of the increased prevalence of both vertebral ankylosing hyperostosis and HLA-B27 in Pima Indians, a possible association between the two was thought to exist. Therefore, histocompatibility testing was done on 44 adult male Pima Indians age 55 or older with classic vertebral ankylosing hyperostosis and 33 age-matched controls. No significant association between any histocompatibility antigen phenotype or haplotype and vertebral ankylosing hyperostosis was found. This disease, therefore, connot be classified with the HLA-B27 associated spondylarthropathies.

Aged↗
PubMed1985

Recognition of a positive MLR within 4 hours using a carrier free electrophoresis system.

Indicator cells--tanned, surface stabilized sheep erythrocytes--were incubated for 1 h in supernatants of 3 h MLCs. Their electrophoretic mobility was measured by an analytical, carrier free electrophoresis system. The change in their mobility compared with an appropriate control was calculated in per cent and correlated with the conventional measured MLR-cpm. The correlation of the two quantities is statistically highly significant (p less than 0.01). Furthermore, the difference of the electrophoretic mobility values of the group of HLA-D-identical and the groups of HLA-D-haploidentical or -different donors is significant beyond the 1% level (p less than 0.0005). Our method enables, therefore recognition of a positive or negative MLC after only 4 h. Typing for HLA-D-determinants seems to be possible. This could be of great importance for histocompatibility testing and organ transplantation.

Carrier Proteins↗
PubMed1986

[Mixed lymphocyte culture as a compatibility test for bone marrow transplantation].

The reactions in mixed lymphocyte culture (MLC) between family members from 35 healthy families and 80 families of potential bone marrow recipients are shown. From the low MLC reactions seen in 128 pairs of HLA identical siblings, the limit of "Negative Reaction in Mixed Lymphocyte Culture" valid for our own laboratory can now be defined. The importance of mixed lymphocyte culture as a histocompatibility test for bone marrow transplantation is discussed.

Bone Marrow Transplantation↗
PubMed1997

Strategies for determining HLA compatibility in related donor bone marrow transplantation.

BACKGROUND: Although HLA identity between donor and recipient is no longer an absolute requirement for bone marrow transplantation, knowledge of the degree of HLA compatibility is necessary for determining the induction and immunosuppression regimen to be used. In cases of related donor transplantation, HLA compatibility may be assessed by defining the HLA phenotypes at the allele level using high-resolution, DNA-based typing methods or by determining the genotypes of the patient and potential donor from the HLA phenotypes, ascertained by low-resolution typing, of their family members. METHODS: We developed an algorithm that can be used to assess the relative costs of these two approaches. We applied population frequencies for HLA-DR alleles to this algorithm to determine at what cost per test ratio for high-resolution:low-resolution testing the costs of the two approaches are equal. RESULTS: In transplants involving a sibling pair who have the same HLA-A, -B, and -DR antigens, these values are 1.16-1.83 for African-Americans and 1.23-1.97 for Caucasians, depending on the relatives available for testing. With a slight increase in the resolution level achieved with DR antigen testing, the range of values becomes 1.10-1.74. We also estimated that the probability that two antigenically identical siblings have identical HLA-DRB1 alleles is >99% for both African-Americans and Caucasians. A review of 615 cases from our transplant program showed that all of 192 pairs of antigenically identical patients and sibling donors were genotypically or allelically identical, indicating that this estimate is valid. CONCLUSIONS: Transplant programs can apply these algorithms to determine the most cost-effective scheme for histocompatibility testing.

Algorithms↗
PubMed1996

Analysis of HLA-DRB1 alleles using PCR-RFLP and PCR-MPH.

In this study we compare the results of HLA-DRB1 genotyping by PCR-RFLP and PCR-MPH. HLA-DR specificities were also performed by LCT. Samples were obtained from 20 Thai patients who were on the waiting list for kidney transplant. DNA was extracted by phenol-chloroform extraction. It was found that the results gave complete agreement with two methods of DNA typing, however, there were 3 discrepancies in assigning serologic DR specificities and DNA subtypes (p = 0.0001) which were due to the cross reactive antibodies and the lack of potent antisera to define proper HLA-DR subtypes by LCT. These PCR techniques can be applied to identify other alleles such as HLA-DPB1 and HLA-DQB1 which will improve the standard histocompatibility testing in the future.

Alleles↗
PubMed1995

[Allogeneic bone marrow transplantation in malignant blood diseases].

Allogeneic bone marrow transplantation has an established role in the treatment of malignant blood diseases. For some disorders it is at the moment the only curative treatment. As the complication risks of this treatment increase with age, the upper age limit for allogeneic transplantation is usually 50 to 60 years. The main indications are acute leukaemias, chronic myeloid leukaemia and myelodysplastic syndromes. Selected patients with chronic lymphatic leukaemia, multiple myeloma and lymphoma are also treated with allogeneic transplantation. The most common intensive conditioning regimen preceding the transplantation consists of total body irradiation and cyclophosphamide. The source of haematopoietic stem cells has routinely been bone marrow, but the number of transplantations with stem cells harvested from blood is rapidly increasing. The proportion of transplantations from unrelated donors is growing. Histocompatibility testing is becoming more precise, which is likely to improve the results of unrelated donor transplantations to the level achieved with sibling donor transplantations.

Bone Marrow Transplantation↗
PubMed1999

New high resolution typing strategy for HLA-A locus alleles based on dye terminator sequencing of haplotypic group-specific PCR-amplicons of exon 2 and exon 3.

In this study, a new sequencing-based typing strategy for the HLA-A locus is presented which involves group-specific separate amplification of exon 2 and 3 of HLA-A alleles in a first step. Conserved HLA-A locus-specific primers of intron 1 or 3 were combined in 10 primer-mixes with group-specific primers hybridizing to the 5'- or 3'-end of exon 3 or 2 for pre-typing of the HLA-A alleles in 14 allelic groups. Maximally four overlapping short amplicons are produced under identical polymerase chain reaction (PCR) conditions with individual separate amplification of exon 2 and exon 3 of the haplotypic alleles in most heterozygous combinations. Time- and money-saving one-directional Big Dye Terminator cycle sequencing is shown to provide reliable high resolution typing of the HLA-A alleles, even in a few cases of two amplicons in one primer reaction mixture. In comparison, to other sequencing-based typing (SBT) techniques the applied typing strategy minimizes the risk of unequal amplification or of drop-outs of one of the haplotypic alleles and allows unequivocal definition of the cis/ trans linkage of polymorphic positions of the complete exon 2 and exon 3 in most heterozygous cells. This also includes detection of new alleles differing in the polymorphic template generating primer annealing sites as well as in unusual combinations of known exon 2 and 3 sequences. With 10 primer sets working under identical conditions for pre-grouping and separate amplification of the haplotypic alleles our SBT procedure also could be implemented in clinical settings of large-scale stem cell donor histocompatibility testing for fast molecular HLA-A matching.

Alleles↗
PubMed1994

[Bone marrow transplantation from donors other than HLA matched siblings for hematological malignancies. Nagoya Bone Marrow Transplantation Group and Tokai Marrow Donor Bank].

One hundred and fourteen patients with hematological malignancies received bone marrow transplantation from donors other than HLA-identical siblings. Sixty-three patients received transplantations from related donors; 20 were phenotypically identical for HLA-A, B, D/DR (RM0). 32 differed at one locus (RM1) and 11 differed at more than one loci (RM2). Fifty-one transplantations were from unrelated donors; 37 were phenotypically identical and mixed lymphocyte culture (MLC) negative (UR0) and 14 were MLC positive (UR1). One hundred and four patients had durable engraftment. Four (RM1(1), RM2(2), UR0(1)) failed to achieve engraftment. In terms of the probability of > or = Grade II acute graft-versus-host disease (GVHD), there was no significant difference among the groups according to HLA disparity (RM0:25%, UR0:33%, UR1:39%, RM1:47%, and RM2:50%). The probability of chronic GVHD was significantly higher in UR0 and UR1 than RM0 (71%, 75% vs 28%, p < 0.05). The disease-free survival at 3 years was 45% (RM0), 50% (RM1) and 42% (UR0). More than 50% of patients other than RM0 died of fatal complications including GVHD within 60 days after grafting. In conclusion, unrelated donor and related donor mismatched at one locus could be selected for marrow graft in the case of the absence of an HLA-matched related donor. However, more advances in post-transplant management and in histocompatibility testing should be required.

Adolescent↗
PubMed1995

Cadaver kidney transplantation in the north Italy transplant program in the nineties.

The most relevant changes which have taken place in the NITp in the nineties include the introduction of HLA-DRB1 matching, the extension of both recipient and donor selection criteria, and an increase in donor procurement and consequently, in transplantation activity. Some of the changes in policy resulted from extensive analysis of our previous experience. For the future years, the NITp has set the following priorities: Consolidate the increase of donor procurement activity registered in the past 15 months by educational campaigns for health workers and the public, and organizational measures aimed at strengthening ICUs, nominating transplant coordinators and introducing a system of reimbursement for organs procured. Improve the quality of results in terms of patient rehabilitation and cost-benefit through: continued evaluation of protocols for patient admission on the waiting lists and careful selection of donors; and prospective use of genomic HLA Class II matching and also consider genomic typing for HLA Class I which is almost a reality. Establish a single pool of patients on the waiting list evaluated according to common protocols with the possibility of performing the transplant in each of the authorized centers in turn, respecting the best HLA match and the local use of organs. Finally, standards for donor treatment, organ procurement, histocompatibility testing and transplantation must be established. For this purpose, accreditation programs which have begun to be applied in Europe seem to be the adequate tool.

Adolescent↗
PubMed1980

Nomenclature for factors of the HLA system, 1980.

This article outlines the decisions made by the WHO nomenclature committee on leukocyte antigens at a meeting held after the 8th International Workshop on Histocompatibility Testing. Particular attention is given to new designations for provisional HLA-B and HLA-DR specificities and the upgrading of certain HLA-D and HLA-DR specificities to full HLA status. The existence of supertypic cross-reacting specificities was confirmed and extended.

HLA Antigens↗
PubMed1984

The improving prognosis after kidney transplantation. New strategies to overcome immunologic rejection.

For the 70,000 patients with end-stage renal disease in the United States, renal transplantation offers the only hope of full recovery from chronic renal failure. However, transplantation has had only limited use, principally because of the risks of graft rejection and immunosuppression. The last ten years have witnessed striking improvements in the survival of patients and grafts resulting from advances in immunologic management, including restricted use of immunosuppression, better histocompatibility testing, HLA matching, blood transfusions, and new drugs for prevention and reversal of transplant rejection. Kidney transplantation now is safe and effective and should be considered for most young and middle-aged adults.

Adrenal Cortex Hormones↗
PubMed2000

Fourier-transform infrared spectroscopy as a tool for detecting early lymphocyte activation: a new approach to histocompatibility matching.

Fourier transform infrared (FT-IR) spectroscopy due to its speed and sensitivity is becoming an increasingly powerful tool in the study of cell composition. We outline the potential of FT-IR microspectroscopy in monitoring mitogenic and cell mediated lymphocyte activation. We demonstrate the potential of FT-IR spectroscopy in histocompatibility testing by showing that significant spectral differences (p < 0.001, for the mean integrated intensity of phosphodiester band located in the 1142-996 cm(-1) region) exist between lymphocyte cocultures from pairs of HLA matched and mismatched volunteers after only 90 min of incubation. The preliminary results indicate that early spectral changes measured are due to HLA differences between individuals, although the relative contribution of class I and class II differences has yet to be determined. FT-IR spectroscopy represents a novel approach to histocompatibility matching and the rapidity and sensitivity of the technique indicates a potential role in matching protocols for clinical use, particularly in allogeneic bone marrow transplantation.

Analysis of Variance↗
PubMed1977

HLA determination in renal transplantation with living donor.

During the period Feb. 1967--Aug. 1976 94 first renal transplantations were performed using living related donors and histocompatibility tests. Eight transplantations were performed on children under 15 years old and four on patients over 50 years old. The rest of the patients were between 15 and 50 years old. No exclusions were performed. 22 patients expired, seven of these with a well functioning graft. The patient survival (P.S.) was 90% at 1 year, 87% at 2 years and 77% at 5 years, the graft survival (G.S.) was 81% at 1 year, 80% at 2 years and 68% at 5 years. The clinical results showed a fairly good correlation with the histocompatibility degree. In the A-B-match groups the 1, 2 resp. 5 years P.S. was 95, 96 resp. 89% and G.S. 90, 90 resp. 89%. In 43% of the patients rejection of variable degree developed. The rejection led to graft loss in 12%.

Adolescent↗
PubMed1990

HLA histocompatibility affects cardiac transplant rejection and may provide one basis for organ allocation.

Prospective human lymphocyte antigen (HLA) typing is not performed for heart transplantation, and the relation between HLA matching and cardiac graft rejection is unclear. Recipient and donor HLA matching were analyzed retrospectively in 51 patients undergoing orthotopic cardiac transplantation. Immunosuppression was based on cyclosporine and prednisone. During the mean follow-up of 34 months (range, 16 to 63 months), the 46 operative survivors had an average of 3.95 rejection episodes (range, zero to 11 episodes). Twenty-one patients had steroid-resistant rejection requiring treatment with polyclonal or monoclonal antithymocyte globulin. Human lymphocyte antigen typing was available for 44 patients, and antigens were grouped in broad specificities. Patients with two or more HLA-A or HLA-B matches had a reduced number of rejection episodes (3/10 versus 19/34) and a lower incidence of steroid-resistant rejection (1/10 versus 18/34; p = 0.01). Inclusion of HLA-DR matches did not alter the findings. There was a strong correlation between the increased frequency of rejection and the incidence of steroid-resistant rejection (p less than 0.0001). Four of six late deaths occurred in patients with steroid-resistant rejection; four were due to acute rejection and two to graft atherosclerosis. Although not currently done, prospective HLA matching is feasible with present typing methods. Our results suggest a rationale for prospective histocompatibility testing in cardiac transplantation with allocation of donor hearts to patients with two or more HLA matches.

Adult↗
PubMed1992

[Indications, problems and future perspectives of bone marrow transplantation in pediatrics].

Bone marrow transplantation (BMT) is an established therapy in pediatric oncology and is increasingly used as curative approach in the treatment of congenital, non-oncologic diseases of the lymphohematopoietic system. There is increasing evidence, however, that BMT can be followed by severe long term effects including neuroendocrine, ophthalmologic, dental and central nervous system abnormalities, particularly in children. Therefore the indication to BMT depends on the results obtained by conventional therapy. Due to the high cure rates of leukemia with conventional therapy BMT is only warranted following relapse except for certain forms with poor prognosis factors. For patients with chronic myelogeneous leukemia, however, BMT is the only chance of cure. In solid tumors the role of BMT is more difficult, because there is no clear evidence that BMT is superior to conventional therapy with regard to long term survival. In severe aplastic anemia, however, the long term results of BMT are clearly better than those obtained by conventional therapy. Other undisputed indications for BMT are severe combined immuno deficiencies and other congenital diseases for which BMT is currently the only curative therapy. Progress with matched unrelated donor transplantations by better histocompatibility testing and more specific immunosuppressive therapy to reduce graft-versus-host disease, still a major problem of allogeneic BMT, as well as the perspectives of gen therapy in the future will offer a chance of cure to many patients without a matched sibling donor.

Anemia, Aplastic↗