Hla typing is a kind of genetic test used to identify certain individual variations in a persons immune system. In the 1960s, this was the sole method of determining tissue types. The mhc major histocompatibility complex contains more than 200 genes which are situated on the short arm of chromosome 6 at 6p21. Nextgeneration sequencing ngs is the latest technology used to perform hla typing, offering better precision at a lower cost than traditional techniques such as ld pcr hosomichi et al. Method open access hla typing from rnaseq sequence. Ashi and the college of american pathology cap established guidelines for quality control and quality assurance for molecular hla typing laboratory methods. Several advantages for establishing hla molecular typing techniques, one of them is hematopoietic stem cell transplantation from an hla matched unrelated donors.
The ssp based hla typing is an economical, rapid, precise, technically simple and reproducible method. Understanding crossmatch as well as hla typing for renal transplantation and applying it in clinical practice is the key step to achieve a successful outcome. Hla typing a comparison of serology and dna techniques. Analysis of human leukocyte antigen hla immunogenetic. Histocompatibility testing methods medical immunology. One of the first methods of tissue typing was through serological typing.
The shortterm survival rate of organ transplantation has been greatly improved since. The human leukocyte antigen hla system or complex is a gene complex encoding the major histocompatibility complex mhc proteins in humans. A novel multiplexed 11 locus hla full gene amplification. A complementdependent cytotoxicity cdc test or microlymphocytotoxicity assay for hla typing has been developed in the 60s. When a patient is put on the transplant list, the hla antigens are listed into a special computer system, along with several other pieces of information. These cellsurface proteins are responsible for the regulation of the immune system in humans.
The information presented, much of it previously unpublished, was written by leading. If the antibodies recognize their epitope on the donors hla then complement activation occurs leads to cell lysis and death, allowing the cells to take up a dye trypan blue. From december 2011 to december 2012, a prospective determination of the hla types of 20 individuals referred to the tissue typing. Tissue typing for transplantation refers to determination of the hla genotypes of both the potential donor s and the recipient.
Dec 24, 2017 contemporary crossmatch methods and human leukocyte antigen hla typing play a pivotal role in improving organ allocation and afford better matches to recipients. The hla antigen is present on the surface of white blood cells. Analysis of human leukocyte antigen hla immunogenetic data. Several molecular techniques have been adapted for the human leukocyte antigen hla typing with their advantages and drawbacks, which greatly improved the feasibility and precision of hla typing. Hla antigens are cell membrane glycoproteins with key roles in the initiation of the immune response. Hla association studies must use typing techniques with high resolution that distinguish the hla subtypes. The challenges of hla allele level typing strategies for a diverse patient population. Hla typing is performed for kidney, bone marrow, pancreas and heart transplants etc. The degree of human leukocyte antigen hla matching between the transplant recipient and the hematopoietic cell graft has a significant impact the outcomes of related and unrelated. Hla typing, is essential for clinical work for example, organ transplantation, immune system research, and biomarker and drug development. Class i typing methods include test such as microcytoxicity for typing a, b, c loci and cellular techniques such as cml for hla dpw typing. Pdf preliminary report of hla dna typing of nigerians.
Hla matching between donor and recipient is desirable for allogenic transplantation. White blood cells form one of the key agents of the human immune system. Different dnabased molecular techniques are used depending on the clinical application. Hla basics human leukocyte antigen hla typing is used to match patients and donors for bone marrow or cord blood transplants. Hla are proteins or markers found on most cells in your body.
Careful interpretation of hla typing and crossmatch tests. The hla gene complex resides on a 3 mbp stretch within chromosome 6p21. The advanced hla typing strategies for hematopoietic stem. The information presented, much of it previously unpublished, was written by leading authorities in the field of transplantation.
Explore hla resources and education to learn more about hla and the search process. The chapters in this book discuss highthroughput methods for hla. Your immune system uses these markers to recognize which cells belong in your body and which do not. The advanced hla typing strategies for hematopoietic stem cell transplantation. Ngs in the hla typing context requires specificity, fidelity, and robustness to work with a wide range of complex dna templates. Pcr primers and oligonucleotide probes can be designed and validated inhouse, or purchased commercially. Hla antigens expressed on the surface of the lymphocytes of the recipient are matched against those from various donors. Education series the hla education series targets a wide range of knowledge levels around the role of hla in cellular transplantation. Your immune system uses these markers to recognize which cells belong in.
The accuracy of these typing techniques is important because organ and tissue transplantation rejection, 1human genetics laboratory, university of madeira, campus of penteada, 9000399 funchal, portugal. Current hla typing techniques use labor and timeintensive methods, such as sequencespecific oligonucleotide probe ssop hybridization 1, pcr amplification with sequencespecific primers. The polymorphic nature of hla alleles was initially found and defined by serologic methods and the hla antigens were given numbers. When a potential donor is identified, the donor is tissue typed and the donors hla. Hla typing provides evidence of tissue compatibility. Frontiers nextgeneration sequencing based hla typing. For a list of the available dna samples with all available hla typing, please view the pdf or excel document.
Hla polymorphism was first identified and characterized using alloantibodies antisera against leukocytes and the microlymphocytotoxicity test was developed for serological typing of hla antigens tissue types in humans choo, 2007. In this technique, a donors blood cells are hla typed by mixing them with serum containing anti hla antibodies. This technique is timeconsuming for hla typing where the number of probes to detect all specificities is very high. Hla typing was initially performed using serological techniques. This communication is an attempt to present the experience and a preliminary report of results over a oneyear period. Future perspectives in hla typing technologies intechopen. Polysolver uses relatively low coverage whole exome sequencing wes data and a subsequent mutation detection pipeline that is based on the inferred alleles to detect mutations in hla genes. Hla testing is performed and test results are kept on file to compare with results from those people who need a donor match. Human leukocyte antigen hla typing and matching be the match.
Careful interpretation of hla typing and crossmatch tests in. The application of molecular techniques to hla typing began around 1987 when the southern blot technique was used to identify restriction fragment length polymorphisms rflps associated. Hla matching in hsct david smillie h principles of hla typing. Enable costeffective and significant growth of donor registries. Oct 25, 2018 nextgeneration sequencing based hla typing.
Ssp depends on dna amplification using group or allelespecific primers and detecting an amplified product of the correct size by gel electrophoresis figure 30. Previously, sequencing an entire hla gene in isolation was achieved through the use of pcrcloning processes, which are lengthy and often problematic. During tissue typing, a number of hla genes should be typed in both the donor and recipient, including hla class i a, b, and c genes, as well as hla class ii drb1, drb3, drb4, drb5, dqa1, dqb1, dpa1, and dpb1 genes. In addition, hla typing and anti hla antibody screening is performed for transfusion support services. Hla antibody screening of recipients hla antibody testing is performed on the recipient to determine if there are any antibodies present that would target the donated organ or tissue. Methods for hla typing are either based on detection of genetic variation in the expressed hla molecules serological typing, or now almost universally, at dna sequence level dna typing. Most highresolution fourdigit hla typing technologies, mainly sequencebased techniques sbt or probebased hybridization techniques, primarily focus on deciphering the sequence of the antigenbinding groove only. Even though this method is still performed in some laboratories today, there are numerous limitations. Hla typing is made more difficult by the fact that the hla region is the most genetically variable region in the human genome. Significant associations have been found between the hla alleles and certain disorders or deficiencies in many disciplines, including rheumatology, nephrology, neurology, endocrinology, gastroenterology, respiratory, ophthalmology. The oragene sample collection product line provides an allinone system for the collection, stabilization and transportation of dna from saliva that will improve access to potential donors, maximize compliance, and meet your downstream hla typing needs. Molecular polymerase chain reaction pcr based hla typing techniques allow the determination of dna sequence variations, either by hybridisation with sequencespecific oligonucleotide probes pcrsso, or by a combination of sequence specific primers pcrssp, or by direct sequencing pcrsbt of pcr product. However, unambiguous hla typing remains challenging. Given the large number of rnaseq profiles in the public domain and our efforts to develop individualized t cellmediated cancer vaccines, which require the knowledge of a patient hla type and hla expression to prioritize target epitopes 1012, we sought to develop an algorithm to utilize the sequence content of rnaseq reads to determine.
The selection of the optimal donor is based on highresolution hla typing. Advancement in computational techniques in population genetics to better handle hla typing ambiguity has improved calculation of hla haplotype frequencies, prediction of allelelevel hla typing for subjects with typing ambiguity in registry matching algorithms, and projection of hla match likelihoods as registries expand. Serology is still the method of choice in many laboratories for low resolution typing of hlaa and hlab discrimination between groups of related alleles only, owed to its simplicity and low cost. Contamination of dna from the previously amplified dna product is an important consideration in molecular hla typing laboratories.
Current publications and web site listing all class i and ii nucleotide sequences and nomenclature. The clinical applications of hla typing most commonly inlcude 1 typing for renal transplantation as one component in the scheme to allocate deceased donor kidneys to individuals on the renal transplant waiting list because of the longterm benefit of using hla matched kidneys, 2 typing for hematopoeitic stem cell transplantation hsc as it. There are various types given to different individuals. Current methods for hla typing define hla alleles and allele groups using dnabased methods. Hla typing bioinformatics tools wes analysis omicx. Dna may be ordered as individual samples, quality control panels, or reference panels. Molecular hla typing methods used in clinical laboratories. Mar 27, 2012 hla typing methods, techniques and results. Hla typing and matching multiple pre and posttransplant factors including patient, disease stage, and transplantrelated aspects can impact transplant outcomes. These techniques provide an opportunity for single allele definition at primary hla typing as opposed to crossreferencing results from different molecular techniques and serological testing. Bellone, in reference module in biomedical sciences, 2014. A number of different dnabased hla typing methods are in everyday use in clinical hla typing laboratories, all of which are based on pcr amplification of target sequences in the hla genes under investigation. A problem in class i typing is the large number of probes required to give. The great search hla typing, donor selection and beyond.
The histocompatibility laboratory performs hla typing for disease association studies such as narcolepsy hla dqb10602, uveitis hla b27, ankylosing spondylitis hla b27 and bird shot retinopathy hla a29 as well as others. Method open access hla typing from rnaseq sequence reads. Purified genomic dna from the bcell lines is also available. Human leukocyte antigen hla typing, utilising the sequencespecific oligonucleotide sso and sequencespecific primer ssp technologies, has been in routine use in many tissue typing laboratories worldwide for more than 20 years since the development of the polymerase chain reaction. Next generation sequencing ngs has been shown to resolve the hla typing ambiguity and simplify hla typing workflows. Hi all, i was performing hla typing for few samples with the use of hlareporter. In the mid1990s, dnabased techniques became more popular and were used to complement serological techniques. Later it was found that antigens with same serologic specificity may have different amino acid sequences. Finding the best donor for a kidney transplant generally means finding a sixantigen match by looking at each of the two alleles at hla a,b, and dr. Handbook of hla typing techniques is an authoritative collection of hla phenotyping and genotyping techniques to be used at the bench level and as a reference. Unfortunately this tool does not include all genes that i would like to analyse. Typing and antibody detection in the laboratory of immunogenetics. Basic biology and clinical applications could not wait for a wellmatched donor.
Serological hla typing hla typing methods were originally based on the. Conventional techniques v nextgeneration sequencing. Contemporary crossmatch methods and human leukocyte antigen hla typing play a pivotal role in improving organ allocation and afford better matches to recipients. This volume explores the rapidly evolving field of hla typing and its use in both the laboratory setting and in silico methods. We use your linkedin profile and activity data to personalize ads and to show you more relevant ads.
Hla typing and its influence on organ transplantation. Class ii typing involves cellular techniques such as. The rapid progress of hla typing techniques has contributed to improving the outcome of haematopoietic stem cell transplantation hsct. Identification of human leukocyte antigen hla patterns in. Enables high precision human leukocyte antigen hla typing even. Serology is still the method of choice in many laboratories for low resolution typing of hla a and hla b discrimination between groups of related alleles only, owed to its simplicity and low cost. Hla typing stands for human leukocyte antigen typing. Handbook of hla typing techniques 1st edition kam m. A sixantigen hla a, hla b, and hla dr match confers a graft survival advantage compared with 0 antigen matches for both cadaveric and live donor transplantation of 10% at 10 years. Human leukocyte antigen hla typing and matching be the. The results of dr ssp typing were far superior and almost 90% of alleles were identified. The process is critical for identifying which people can safely donate bone marrow, cord blood, or an organ to a person who needs a transplant. It was a relief to know that the use of randomlv matched donors was not going to result in an intolerable pealty.
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