Laboratory Testing for Compatibility in Transplantation

LABORATORY TESTING FOR COMPATIBILITY IN TRANSPLANTATION

HLA Typing

Histocompatibility between the donor and recipient is primarily defi ned by  genes of the MHC. MHC is a general term for the gene complex encoding the HLA, and hence the two acronyms are used interchangeably in humans. Historically, HLA typing was performed by serologic evaluation, using a panel of anti-HLA antibody containing sera derived from multiparous women. Most clinical HLA laboratories now use DNA-based HLA typing methods. By using the polymerase chain reaction (PCR), polymorphic resi-dues of both class I and class II HLA or MHC molecules can be amplified using primers that bind to the conserved regions of the gene. The precise nucleic acid sequence of the HLA alleles can be accu-rately determined using sequence-based typing. More commonly, partial ampli-fication of polymorphic residues using sequence-specific primer pairs (PCR-SSP or PCR-SSOP) can identify HLA alleles. Depending on the DNA probe, PCR-based HLA typing is described as either “intermediate-resolution” (beyond the serologic level, but short of the allelic subtype level) or “high-resolution” (allele- or allelic subtype–level typing). The resolution of the HLA typing defines the level of HLA mismatch. Low-resolu-tion serologic typing can identify antigen mismatch (HLA-A2 vs. -A11), whereas high-resolution typing can identify an allele- or allelic subtype-level mismatch (HLA-A*0201 vs. -A*0205).

ABO Blood Typing

ABO blood typing is one of the most important tests in human solid organ transplantation.

ABO antigens are primarily expressed by erythrocytes but also can be expressed by platelets and glandular epithelial and endothelial cells. Incompatibility  occurs when recipients lacking a certain  blood type produce IgM antibodies against that antigen and cause subsequent activation of complement and lysis of transfused incompatible red blood cells. Because ABO antigens are also expressed by endothelial cells, incompatibilities can lead to hyperacute vascular rejection.

In contrast to solid organ transplan-tation, ABO matching does not directly affect donor selection for allogeneic HSCT. Because inheritance of blood group anti-gens is independent of HLA antigens, fully HLA-matched donor-recipient pairs are often ABO incompatible. ABO incompat-ibility requires red blood cell depletion of a stem cell allograft but does not affect myeloid or megakaryocytic engraftment, graft rejection, or graft-versus-host disease (GvHD). Upon engraftment, the recipient transitions to the donor’s ABO blood type. During the conversion from recipient to donor blood type, transient hemolysis may occur.

Screening for Preformed Antibodies

Patients awaiting cadaveric organs un-dergo screening of their blood for pre-formed antibodies against HLA molecules. These antibodies may be produced by pregnancy, prior blood transfusions, or prior organ transplantation. An aliquot of the patient’s serum is mixed in separate wells with cells of at least forty donors assumed to be representative of the donor population. Complement-mediated lysis or flow cytometry with fluorescent-labeled secondary antibodies to human IgG quantifies the number of reactive cells as a proportion of the number of panel cells (percent reactive antibody, or PRA). Because transplantation across a posi-tive cross match can cause hyperacute or acute rejection, patients with high PRAs typically experience longer waiting times on the cadaveric organ donor list. Many centers are therefore currently exploring strategies to remove these antibodies, by plasmapheresis, for example, either preop-eratively or perioperatively.

Cross Matching

In solid organ transplants, cross matching is usually done after a potential donor is identified. The recipient’s serum is tested for reactivity to the donor’s lymphocytes using the previously mentioned comple-ment-mediated lysis or flow cytometric assays. A negative cross match means that there is no recipient antibody that is reac-tive with donor cells or graft. A positive cross match portends severe rejection if the donor organ is transplanted.