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HLA and PRA Testing

Blood Group Matching

In order to match a potential recipient to a donor organ one first must ascertain that the blood groups are compatible. The following chart shows which particular blood groups can donate to, or receive from, the other blood groups. (Rh factor does not enter into tissue typing decisions.*)

The chart shows the possible combinations of tissue compatibility, not necessarily the practical application of blood group matching. “For reasons of fairness, organs are allocated primarily to their own blood group. Otherwise, the O patients would only have access to a fraction of the organs, while AB patients would have access to all organs. Nevertheless, there are some inequities in the waiting times on particular blood group lists.” (3)

Finally, …[blood group matching] does not seem to make much of a difference in the case of liver transplants; the reason for this is unclear. [It]…is known that one can use `blood group incompatible' livers (an A liver in a B patient) with success rates almost as good as blood-group identical livers. [The choice]…still [is the] use [of] blood group identical livers when at all possible, because the success rate is higher overall. The allocation schemes for organs takes these principles into account. (3)

Certain blood group combinations carry strict prohibitions, but the choice of compatible blood group matches is less rigid.

HLA and PRA: Transplant Tissue Typing

Earlier, we discussed the Major Histocompatibility Complex and the Class I and Class II antigens. The potential match between the recipient's tissues and the donor's tissues uses a method call Human Leukocyte Antigen testing, or HLA.

HLA testing either includes or excludes a particular recipient and a particular donor as a transplant match. HLA looks at six markers, antigens that are carried on the surface of the body's leukocytes. The Class I antigen markers are designated HLA-A, HLA-B, and HLA-C; the Class II antigen markers are designated HLA-DP, HLA-DQ, and HLA-DR. The Class I antigens directly activate the T-8, or killer T-cells (cytotoxic T-cells), while the Class II antigens principally activate the helper T-4 cells, macrophages, and B-cells. Since these six HLA antigens are carried on the surface of the cells, they are highly reactive with the B- and T-cells, and therefore, the closest possible match is required for the best results in transplantation.

The number of specific proteins encoded by these 6 antigens (actually, the two sets of three antigens) is quite large. One source gives “24 specificities encoded by HLA-A, 52 by HLA-B, and 11 by HLA-C [and]…there are at least 20 specificities encoded for HLA-DR, nine for HLA-DQ, and six for HLA-DP.” (4) Estimates of the total number of possible HLA combinations vary between 10,000 and 13,700.

Antigen/Peptide Complex of MHC

To perform the HLA test, samples of known HLA antigens are incubated with samples of serum (containing the subject's antibodies); a positive reaction indicates the presence of that specific antigen on the subject's cells. A close match between recipient and donor means that these markers of "self" are similar or identical, making the immune system less apt to attack the transplant.

The Panel Reactive Antibody test (sometimes referred to as Percent Reactive Antibody, since the result is expressed as a percentage), or PRA, also tests histocompatibility. This test combines the subject's serum with samples of cells (which contain antigens) taken from up to 60 different individuals. The combination again shows if the subject has acquired antibodies (for example, from a blood transfusion) to a spectrum of antigens. Possible antibody/antigen mismatches are preliminarily screened out. By counting the number of reacted cells in a given sample, the PRA test also shows how strongly the antibodies react with the antigen. If 50 or 100 cells react, then that sample is 50% reactive. The results of this test, then, show which antigens react with a subject's antibodies, and how severely they react. Obviously, in transplantation better matches are indicated by fewer reactive samples, and less severely reactive samples.

After the HLA test identifies a potential donor/recipient match, and the PRA testing eliminates a broad cross-section of antibody/antigen mismatches, a crossmatch is performed. The crossmatch combines the donor's cells with the recipient's serum and vice-versa, to verify that antigens and antibodies from both donor and recipient are non-reactive, or at least minimally reactive.

These three tests, then, show us what the donor's tissues and the recipient's body consider “self” and “non-self”, and they show us what antigens and antibodies should be avoided in a potential match. Since the HLA markers are inherited from one's parents, the markers remain constant; the lab must perform this test only once. However, since antibodies can arise or subside, the PRA test should be repeated at least monthly; if the potential recipient receives a blood transfusion, the test should be repeated in no more than 10-14 days.

Living Donors, Cadaveric Donors, and HLA

Parents provide one's specific combination of HLA markers. Of the six markers present on a child's leukocytes, a group of three derives from the father, the other group of three derives from the mother. Since the child inherits each half—each haplotype—from a parent, a child-parent match is almost always a one-haplotype match (i.e., 3 of 6 HLA match). (A theoretical possibility would be that both parents coincidentally share an identical haplotype; this could make the child an identical match to one parent, and a one-haplotype match to the other. However, the vast number of different HLA antigens considered, the chance of this happening seems quite rare.)

Since one's siblings also combine some of the same six maternal and six paternal antigens (although not necessarily the same three antigens from each parent), they can offer a match ranging from a zero of six antigen match, up to a six of six antigen match. The selection of antigens is an individual occurrence with each child, so while one can calculate the statistical odds of a perfect match, there is no guarantee that it will occur, no matter how many siblings one may have. Aunts, uncles, and other relatives can make possible matches, as can totally unrelated individuals.

Cadaveric donors draw from a more random gene pool, at least, compared to that of a recipient and family. Still cadaveric donors provide the largest source for kidneys, the only source for heart transplants, and very close matches are quite common. A six-of-six HLA match with no PRA reactivity is the best possible match. Five-of-six matches, and four-of-six matches are more common and are considered quite good matches. Depending upon the rarity of the blood group and the specific HLA antigens, matches that are theoretically “less perfect” are frequently made, and with modern immunosuppressant drug therapy, are very often long-term successes.

Living related donor transplants still have an edge in terms of the longevity of graft survival. This fact is generally thought to have as much to do with the physical and biochemical condition of the organ at the time of transplant as with the exactitude of the HLA match:

A wide variety of powerful vascular mediators are released in the presence of severe brain injury. As a consequence of this 'catecholamine storm,' hearts and lungs from cadaver donors may manifest such severe deterioration in function that they become too damaged to transplant. All cadaver organs sustain this type of injury, some to a greater degree than others. For example, hearts damaged in this way manifest lesions, such as contraction band necrosis, and kidneys manifest acute tubular necrosis. In fact, in kidneys, this effect is stronger than the effect of HLA matching as evidenced by the data that shows that the outcomes from kidneys from spousal donors are superior to outcomes from cadaver donors with the same degree of HLA matching. (5)

Still, with improving drug therapy, cadaveric donors are gaining ground relative to living-donor graft longevity.

Tissue typing is a complex process, and the foregoing is just an outline of the major steps in recipient/donor matching. A large portion of this matching must be done accurately and rapidly in the last hours before transplantation (speed being essential in the case of cadaveric donors). Fortunately the technology exists to make possible quick, precise tissue typing for optimal transplant results.

*The reason that Rh factor does not affect tissue typing is that Rh factor affects only erythrocytes, or red cells, whereas HLA typing is dependent upon leukocytes, or white cells, and their properties.