Future Perspectives in HLA Typing Technologies

Abstract

A wide range of malignant and nonmalignant diseases require hematopoietic stem cell transplantation (HSCT) as last resort therapeutic approach. Graft versus host disease (GVHD), which is one of the major causes of transplant-related mortality, is minimized whenever increased matching of human leukocyte antigens (HLAs) between donor and recipient is present. Suitable donor selection is determined with the utilization of HLA typing. HLAs are highly polymorphic glycoproteins encoded by a region of genes known as the major histocompatibility complex (MHC). Their biological function is to present antigenic peptides to T lymphocytes. However, they also play important role in HSCT acceptance/rejection. During the previous years, various techniques have been acquired in order to better characterize the HLA profile of transplant donors and recipients. This effort is particularly challenging due to MHC size, but most importantly due to high sequence variability in specific regions of the respective genetic loci, between individuals. Initially, HLA typing was performed using serological typing, hybridization techniques, and restriction fragment length polymorphism (RFLP) approaches. Later on, polymerase chain reaction (PCR) based techniques and direct sequencing (dideoxy-based Sanger sequencing) capillary electrophoretic analyses arose. Nowadays, 2nd and 3rd generation sequencing (NGS) technologies show great potential in effectively identifying these polymorphic regions.

Tissue typing for kidney transplantation for the general nephrologist

“…First‐generation molecular typing is based on DNA amplification by polymerase chain reaction (PCR) (Sequence‐Specific Oligonucleotide and Sequence‐Specific Primers techniques) with subsequent sequencing of the amplified gene product to define HLA type . Next‐ or second‐generation sequencing also uses PCR gene amplification after first digesting the DNA into short fragments and amplifying the fragments from multiple points to generate vast numbers of DNA copies that are read and virtually reconstructed using computer algorithms . This improves the accuracy of the typing.…”

“…The techniques used are complex but in simple terms make use of panels of ‘barcoded’ molecules that can bind directly to the HLA gene. The molecules which bind the individual's DNA have their barcode read to determine the HLA type . These techniques are highly efficient requiring only a very small DNA sample.…”