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Saudi Journal of Kidney Diseases and Transplantation
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REVIEW ARTICLE Table of Contents   
Year : 1997  |  Volume : 8  |  Issue : 2  |  Page : 138-144
Consensus and Controversies on HLA Matching and Crossmatching in Transplantation

Director, Laboratory and Blood Bank, King Saud Hospital, Unaizah, Al-Qassim, Saudi Arabia

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Transplantation of organs is becoming increasingly successful. What was once an experimental and life-saving emergency procedure has now been transformed into a life-enhancing and technologically advanced form of therapy. Histocompatibility testing between the donor and recipient consist of identification of HLA antigens known as tissue typing, and testing for pre-formed antibodies in the recipient against the donor HLA antigens, known as crossmatching. The importance of matching of HLA antigens has been debated ever since the first successful renal transplant was performed in 1954. Powerful and newer immunosuppressive drugs have not been able to establish an operational state of long-term organ tolerance. In this article, an attempt is made to show the clear and obvious effect of HLA antigen matching on the long-term graft survival. Also, the merits and demerits of different crossmatching methods and their proper interpretation and application to improve the overall graft survival are discussed.

Keywords: Tissue typing, Crossmatching, Long-term graft survival, Immunosuppression, Transplantation.

How to cite this article:
Qureshi B H. Consensus and Controversies on HLA Matching and Crossmatching in Transplantation. Saudi J Kidney Dis Transpl 1997;8:138-44

How to cite this URL:
Qureshi B H. Consensus and Controversies on HLA Matching and Crossmatching in Transplantation. Saudi J Kidney Dis Transpl [serial online] 1997 [cited 2021 May 5];8:138-44. Available from: https://www.sjkdt.org/text.asp?1997/8/2/138/39387
Transplantation has come of age and its future looks bright. It is predicted that by the year 2000, a large number of surgeries will be replacement surgery. Transplanting the lung, heart, liver and kidneys have now become common practice. Small bowel and islet cell transplantations are other examples of the advances and pioneering efforts made in the field of transplantation. The era of organ transplantation has crossed 25 years and the transplant surgeons have been constantly trying to break the immunological barrier by developing and introducing powerful immunosuppressive drugs. Modern immunosuppressive drugs have played an important role in the success of trans­plantation. However, histocompatibility and immunology also play important roles in transplantation. Histocompatibility tests including matching for HLA antigens and crossmatching remain an integral part of pre-transplant work-up.

   HLA Matching Top

For over 25 years now, it has been difficult to establish the relevance of HLA matching in clinical transplantation. This was evident from a large scale study in 1970 which suggested that HLA incompa­tibilities can be successfully over-ridden by immunosuppression [1] . This observation has remained unchanged over the ensuing years, and success rates for HLA-incom­patible transplants have increased steadily during the 20 years since 1972 [2] . Heart and liver transplants are being performed today with no regard to matching, and they have approximately the same graft survival rates as kidney transplants, for which attempts are made to match the donors and recipients [3],[4] .

The above factors not withstanding, HLA matching seems of be an important factor as evident by the observation that the highest graft survival rates are achieved when both HLA chromosomes are matched, as in HLA-identical siblings [2] . Also, one-­haplotype-matched parental donor organ grafts have clear advantage over two­-haplotype-mismatched cadaver-donor organ transplants [2] . The effect of HLA matching has been seen most profoundly on long­term graft survival; the half-life of grafts from HLA-identical siblings is 20 years, from parent donors, is 12 years and from cadaver donors the half-life is eight years [2] . Experience with cyclosporine-A (CSA) over the past 10 years has shown that this drug has probably little effect in governing the long-term survival of the grafts with kidney transplants performed 20 years ago and those performed in the CSA era having similar half lives [2] . A positive effect of HLA matching on kidney graft outcome has been clearly documented in reports from two large studies of renal transplant data namely the UCLA transplant registry which has collected data on 106,000 transplants and the collaborative transplant study (CTS), Heidelberg, Germany with data on 107,500 renal transplants [5],[6],[7] . Both these studies indicate that the main factor that improves long-term (10 years and above) renal allograft survival is the donor-recipient matching for HLA antigens [Table - 1]. Also, it has been suggested that matching for the splits (sub­types) of HLA antigens may be even more predictive of graft outcome than simple matching of the generic HLA antigen [5],[6],[7] .

Despite the dramatic improvement achieved in 1-year graft survival, however, the rate of graft loss due to chronic rejection remains essentially unchanged i.e., half of cadaver grafts are still lost by 7.8 years compared with 7.3 years in 1978 [7] . Thus, the use of CSA has not established an operational state of long-term organ tolerance. Opelz has analyzed pooled data from transplant-centers world-wide and observed that long­term allograft survival is inversely related to the degree of HLA A,B and DR mismatch in CSA treated renal transplant recipients [Table - 2]. There was a constant risk of loosing a renal allograft beyond the first year post-transplant. Also, the allograft half-lives were substantially higher with better degree of matching (ranging from 6.8 years for six mismatches to 11.3 years with no mismatches) [5] . Overall, cadaver allograft survival in this group was projected as 40% at 10 years with a graft half-life of 8.8 years. Simultaneous analysis of 5,262 one haplotype matched living-related allografts treated with CSA and 973 HLA-identical allografts showed 10-year projected survival rates of 52% and 73% and graft half lives of 11.9 and 23.6 years respectively.

Regardless of the advances made in the field of immunosuppressive therapy, it is clear that less, rather than more, immuno­suppression is far more desirable. Under­standably, it is the lower levels of immuno­suppression administered to HLA-identical siblings that results in transplants with less complications and longer survival with a half-life of over 20 years. Post-transplant complications are almost always traceable to immunosuppressive therapy. Thus, it is obvious that a graft that requires less. immunosuppression is preferable to a graft requiring more intensive treatment. Good matching for HLA antigens might help in this regard.

Recent experience has shown that many factors affect the outcome of kidney trans­plantation. It seems that the effect of the transplant center (center effect) and HLA matching are the strongest factors [8] . Several medical centers have reported remarkable long-term survival rates [9],[10] . A number of social factors, many of which are difficult to change contribute to this so-called center effect. However, there is evidence now that histocompatibility is a factor that can override the center effect, as demonstrated by the high success rates achieved at all centers with transplants involving a HLA-­identical donor [11] . Therefore, it appears that if the match is good, all centers can have comparable good results.

In the USA, more than 10,000 kidney transplants are performed annually, with cadaver allografts accounting for approxi­mately 80% of them. It is estimated that, if all the available kidneys were shared in the USA nationally, only 25% of all waiting patients can be transplanted with kidneys with no HLA-B or DR mismatches [7] . Therefore, although perfect HLA matching is a good way to improve the transplant outcome, it is not always easy to obtain such a perfect match. Also, in heart trans­plantations, distribution of hearts based on HLA matching is impractical although retrospective analysis indicates an improve­ment in 3-year graft survival when there were less than 2 HLA-B-DR mismatches compared to grafts with greater than 2 mismatches, 83% to 70% survival respectively [7] . For liver transplantation, better HLA­matched livers are associated with fewer rejection episodes but, paradoxically, liver graft survival results have shown no advantage from HLA matching [7] . The value of HLA matching for liver transplant­ation and other organs such as the pancreas must await the collection of more data. Preliminary data for corneal transplantation show that patients with previously rejected grafts will benefit from a well matched transplant [7] .

Tremendous advances have been made even in the procedural aspects of tissue­typing in recent years. Newer techniques and high quality reagents are giving good quality results and have reduced the time and cost of the procedure. Molecular biology technique such as polymerase chain reaction, SSP and SCO sequence-specific oligo­nucleotides (SSO) are available to rapidly identify the class II genes accurately. Similarly, the monoclonal antibody techno­logy is being applied for the procurement of high quality reagents in large quantities for the microcytotoxicity assays [7] .

   Cross Matching Top


A negative crossmatch is an absolute requirement before renal transplantation and there are several techniques available for detecting antibodies against donor histocompatibility antigens. The transplant physicians must select the techniques which are adequate by way of sensitivity and specificity.

The most important reason for performing a crossmatch before transplantation is to detect pre-formed antibodies that may cause hyperacute rejection of the graft. For many years, the basic NIH and the Amos modified versions of the complement mediated cytotoxicity technique have been widely used for this purpose [12],[13] . More recently, the objective of the crossmatch for renal transplantation has evolved to also include detecting antibodies that represent various degrees of risk for graft loss [14],[15],[16] . However, the association between crossmatch results and risk of graft loss is not as strong as between crossmatch results and hyperacute rejection. Another problem is that patients who have non-HLA anti­bodies may be misclassified as allo-sensitized, especially when low affinity antibodies are not removed by repeated washings. Since these antibodies are not harmful to the graft, a decision to not transplant unfairly discriminates against these patients. A number of techniques are now being used for crossmatching with different laboratories using different techniques [17] , a fact that may be a source of confusion for those not thoroughly familiar with this subject.

The following guidelines describe the merits and limitations of the different tests and provide recommendations based on information currently available. The different techniques available are:

a) Direct complement dependent cytotoxicity (CDC) techniques (for complement fixing antibodies);

Basic NIH technique, no wash

Amos modified (wash) technique

Prolonged incubation technique

b) Indirect or binding techniques;

Anti-human globulin (AHG)

Flow cytometry.

Current American Society of Histocompa­tibility and Immunogenetics (ASHI) standards state that the tests used should be more sensitive than the basic NIH, no wash technique [18] . This can be achieved by the Amos modified and prolonged incubation technique as well as the AHG and flow cytometry, listed in order from the least to the most sensitive [19],[20] . The prolonged incubation technique improves sensitivity but delays reporting results. AHG is fast and sensitive whereas flow cytometry can provide the fastest turn-around time and greatest sensitivity for laboratories with the equipment and expertise to perform this test [21],[22] .

Because of the positive correlation between a positive T cell CDC test and the incidence of hyperacute rejection, detection of anti-HLA antibodies with any of the direct CDC techniques usually contra­indicates transplantation [23],[24],[25] . However, the interpretation of a positive indirect technique, either AHG or flow cytometry, is different; there is a higher risk for graft loss, especially in re-transplant or sensitized patients, but good outcome can be expected in a proportion of patients [26],[27],[28],[29],[30],[31] . Graft outcome in these patients is, in all pro­bability, also dependent on center-specific factors (patient selection, immunosuppression, follow-up strategies, etc.). It follows that the use of crossmatch results to make a decision about transplantation can be different, within certain limits, from center to center.

   HLA Versus Non-HLA Antibodies Top

Some patients develop antibodies directed against non-HLA antigens that often produce a positive CDC crossmatch. These antibodies are not harmful to the graft [14],[15],[16] . Sometimes, these antibodies react with autologous lymphocytes and are categorized as auto-antibodies. HLA specificity can be confirmed by platelet adsorption, inhibition by purified HLA or inhibition by monoclonal antibodies [32] . However, these techniques may be difficult to apply within the time constrains of the crossmatch. Non­HLA antibodies are often of low affinity and are removed when CDC techniques with four washing steps are used [33] . Non-­HLA antibodies are also often IgM. Since IgM activity is abolished by reducing agents such as Dithiotrietol (DTT), these antibodies typically produce a positive CDC result in the absence of DTT but become negative in the presence of DTT [34],[35] . Routine use of DTT has been found to be of practical value in distingui­shing potentially harmful IgG anti-HLA antibodies from the inconsequential IgM anti-non-HLA antibodies [34] . Thus, the outcome of transplants performed across a positive CDC crossmatch that becomes negative in the presence of DTT is similar to transplants with completely negative crossmatch results [35],[36],[37] . However, DTT may reduce potentially harmful anti-­HLA IgM antibodies [15] and can, under certain circumstances, inhibit IgG anti­bodies, leading to an erroneous interpre­tation of the results [37] . Ideally, the crossmatch should be able to distinguish HLA from non-HLA antibodies directly. Until practical approaches to achieve this goal are developed, laboratories should make an effort to assess HLA specificity indirectly. Appropriate options include multiple washing steps, DTT, and flow cytometry. The interpretation of the results must take into account the prior sensiti­zation status of the patient, known presence of auto-antibodies, prior transplants or mismatched donor HLA antigens.

In the past, a positive CDC crossmatch with any prior specimen was considered a contraindication for subsequent transplant­ation in that patient under the assumption that a prior response to donor antigens represented a risk for graft loss [38] . This issue is controversial at the present time. A number of reports have shown that good graft survival can be achieved under such circumstances [39],[40],[41] , while other studies have reported a higher probability of graft loss in these patients [35],[42] . The significance of a past positive, current negative, cross-match may be related to the nature of the allo-antigen exposure of individual patients. Patients who develop transient antibodies after blood transfusions may represent a group with good prognosis, whereas previously transplanted or long­standing sensitized patients may represent a high-risk group [3] .

Some centers have established time limits (months or years before the date of cross­match) to determine when results with a past specimen should be disregarded. However, there is little data to objectively support any such cut-off time. It has been suggested that a past positive crossmatch due to IgG anti-class I antibodies, is associated with poor graft survival [28] . However, the published information on this issue is still insufficient for valid generali­zations. Until more definitive data become available, it may be legitimate to disregard a positive historic crossmatch in a patient who had a transiently positive panel reactive antibodies with the crossmatch being clearly negative. However, it may be advisable not to transplant a patient who has other immunologic risk factors such as a previous transplant or a current positive flow crossmatch.

   The B Cell Crossmatch Top

A number of reports have documented that anti-HLA-DR and DQ antibodies, especially in high titers (> 1:4 or > 1:8), may cause hyper acute rejection of a renal allograft [43],[44],[45] . Although, hyperacute rejection may not always occur, such antibodies certainly represent a definite risk [46] . In the absence of hyperacute rejection, probably in cases with low-titer antibodies, no clear correlation has been established between a positive B cell crossmatch and graft survival [46] . This conclusion is valid only if anti-HLA Class I antibodies are ruled out as the ones causing the positive B cell crossmatch, as graft survival in these cases may be decreased [47],[48] .

The association between B cell antibodies and hyperacute rejection has been observed both in first transplant and re-transplant patients [43],[44],[45] . Therefore, a B cell cross­match should be performed routinely to identify patients with high-titer B cell antibodies. Centers that screen for pre­formed B cell antibodies may restrict the B cell cross-match to only those patients with such antibodies. The interpretation of the results, as well as the definition of high titer, require that individual laboratories establish specific criteria to guide the decision making process.

   Summary Top

a. AHG and/or flow cytometry are very sensitive techniques to detect clinically relevant antibodies. The information these tests provide complements that of the direct CDC assays and facilitates the decision making process at the time that an organ donor becomes available for a particular patient.

b. Laboratories should be able to identify a positive crossmatch that is caused by non-­HLA antibodies. A careful review of the past antibody history of the patient, and the use of multiple wash steps, DTT, flow cytometry and analogous crossmatch, alone or in combination, are appropriate avenues to achieve this objective.

c. The rare patients with high-titer anti­donor B cell antibodies are at risk of hyperacute rejection. Although this compli­cation is not frequent, it can only be avoided by performing the B cell cross-match routinely. A high titer is usually defined as > 1:4 but, because of the limited precision of the CDC assays, significant variations are to be expected among different laboratories.

d. The interpretation of a positive reaction other than by a direct CDC technique, requires detailed knowledge of the test performance, potential technical problems, expected correlation between the different tests, transplantation immunology (prior sensitization, HLA matching etc.), and up-to-date information about correlation with graft outcome. Incorrect interpretation of crossmatch results may put a patient unnecessarily at extra risk for graft loss, or it may deny a patient the opportunity for transplantation.

   Acknowledgment Top

The author acknowledges Ms. Rowena M. Omandap, Medical Secretary for typing this Manuscript.

   References Top

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3.Kriett JM, Tarazi RY, Kaye MP. The registry of the international society for heart transplantation in: Terasaki PI, ed. Clinical transplants 1990. L.A. UCLA tissue typing laboratory 1991;21:7.  Back to cited text no. 3    
4.Belle SH, Detre KM, Beringer KC, Murphy JB, Vaughn WK. Liver transplantation in the USA: 1988 to 1989, in Terasaki PI, ed. Clinical transplant 1990 L.A. UCLA tissue typing laboratory 1991;11:9.  Back to cited text no. 4    
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19.Johnson AH, Rossen RD, Butler EW. Detection of allo-antibodies using a sensitive antiglobulin microcytotoxicity test, identification of low levels of preformed antibodies, in accelerated allograft rejection. Tissue Antigens 1972;2:215.  Back to cited text no. 19    
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23.Pate R, Terasaki PI. Significance of the positive crossmatch test in kidney transplantation. N Eng J Med 1969;280:735-9.  Back to cited text no. 23    
24.Kissmeger N, Dsens F, Peterson VP, et al. Hyperacute rejection of kidney allografts associated with existing humeral antibodies against donor cells. Lancet 1966;1:662.  Back to cited text no. 24    
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38.Goeken NE and the clinical affairs committee. Outcome of renal transplantation following a positive crossmatching with historical sera. Human Immunol 1985;14:77.  Back to cited text no. 38    
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40.Matas AJ, Nehlsen-Cannarella S, Tellis VA, Kuemmel P, Soberman R, Veith FJ. Successful kidney transplantation with current-sera-negative/historical-sera­positive T cell crossmatch. Transplantation 1984;37:111-2.  Back to cited text no. 40  [PUBMED]  
41.Kerman RH, Flechner SM, Van Buren CT, Lorber MI, Kahan BD. Successful transplantation of cyclosporine-treated allograft recipients with serollogically positive historical, but negative preoperative, donor crossmatches. Transplantation 1985;40:615-9.  Back to cited text no. 41  [PUBMED]  
42.Taylor CJ, Chapman JR, Ting A, Morris PJ. Characterization of lymphocytotoxic antibodies causing a positive crossmatch in renal transplantation. Relationship to primary and regraft outcome. Transplantation 1989;48:953-8.  Back to cited text no. 42    
43.Mohanakumar T, Rhodes C, Mendez-Picon G, Goldman M, Moneure C, Lee H. Renal allograft rejection associated with presensitization to HLA- DR antigens. Transplantation 1981;31:93-5.  Back to cited text no. 43    
44.Ahern AT, Artruc SB, DellaPelle P, et al. Hyperacute rejection of HLA-AB identical renal allografts associated with B lymphocyte and endothelial reactive antibodies. Transplantation 1982;33:103-6.  Back to cited text no. 44  [PUBMED]  
45.Scornik JC, LeFor WM, Cicciarelli JC, et al. Hyperacute and acute kidney graft rejection due to antibodies against B cells. Transplantation 1992;54:6f-4.  Back to cited text no. 45    
46.Taylor CJ, Chapman JR, Fuggle SV, Ting A, Morris PJ. A positive B cell crossmatch due to IgG Anti-HLA-DQ antibody present at the time of transplantation in a successful renal allograft. Tissue Antigens 1987;30:104-12.  Back to cited text no. 46    
47.Karuppan SS, Lindholm A, Moller E. Characterization and significance of donor­reactive B cell antibodies in current sera of kidney transplant patients. Transplantation 1990;49:510-5.  Back to cited text no. 47    
48.Phelan Dl, Rodey GE, Flye MW, Hanto DW, Anderson CB, Mohanakumar T. Positive B cell crossmatches: specificity of antibody and graft outcome. Transplant Proc 1989;21:687-8.  Back to cited text no. 48  [PUBMED]  

Correspondence Address:
B H Qureshi
Director, laboratory and Blood Bank, King Saud Hospital, Unaizah, P.O. Box 981, Al-Qassim
Saudi Arabia
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PMID: 18417793

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