B. Susskind

Correlation of ELISA-detected IgG and IgA anti-HLA antibodies in pretransplant sera with renal allograft rejection.

The present study compared the occurrence of rejection episodes during the first twelve posttransplant (Tx) months and the 1-, 2-, and 3-year graft survivals among recipients stratified by the percent panel reactive antibody (% PRA) of pre-Tx sera as detected using either an antihuman globulin determined PRA (AHG-% PRA) or an ELISA methodology detecting IgG reactive against soluble HLA class I antigens (% PRA-STAT). There was a significant correlation between AHG-PRA greater than or equal to 10% and a PRA-STAT greater than or equal to 10% (P<0.001). However, among 200 sera displaying an AHG-PRA greater than or equal to 10% (mean 57 +/- 2l%), only 69% (138/200) displayed a PRA-STAT greater than or equal to 10%. With further study the discrepant finding, of 62 sera that were AHG-PRA greater than or equal to 10% but PRA-STAT <10%, was due to the presence of IgM and/or IgG non-MHC reactivity. In contrast, among 293 sera displaying an AHG-PRA < 100% (mean 3 +/- 2%), 15% (43/293) displayed a PRA-STAT greater than or equal to 10%. There was no correlation between AHG-% PRA and rejection episodes occurring during the first twelve post Tx months. In contrast, however, there was a highly significant correlation between PRA-STAT greater than or equal to 10% and the occurrence of rejection episodes during the first twelve post-Tx months (P < 0.001). Patients with PRA-STAT greater than of equal to 10% experienced a 70% rejection frequency compared with the 35% rejection frequency for patients with PRA-STAT sera < 10% (P<0.001). A significant correlation was observed between the presence of IgG-1 and rejection (P<0.01) but not IgG-subclasses 2, 3, or 4. Of particular interest was the observation in 11 patients that the presence of ELISA-detected IgA anti-HLA class I antigen (ELISA-IgA PRA greater than or equal to 10%) was associated with a significantly reduced rejection risk compared with sera where only PRA-STAT greater than or equal to 10% was present (27% vs. 70% incidence of rejection episodes, P<0.01). Finally, patients displaying pretransplant PRA-STAT results < 10% experienced significantly improved l-, 2-, and 3- year graft survivals of 85% vs. 74%, 82% vs. 70% and 81% vs. 67%, respectively (P<0.01 for each time point), compared with patients displaying PRA-STAT results greater than or equal to 10%. These data suggest that the use of the ELISA methodology to detect IgG reactivity against soluble HLA class I antigens (PRA-STAT) may allow for the determination of a more clinically informative % PRA than the AHG-% PRA. Moreover, the presence of ELISA-detected IgA anti-HLA may act to inhibit rejection mechanisms associated with ELISA-detected IgG anti-HLA greater than or equal to 10%.

Flow cytometry-detected IgG is not a contraindication to renal transplantation : IgM may be beneficial to outcome

BACKGROUND At our transplant center, primary recipients of either a haplo-identical (haplo-ID) living related (LRD) or a cadaveric (CAD) donor renal allograft are transplanted after a negative donor-specific IgG anti-human globulin (AHG) cross-match (XM). Testing included the historically highest panel-reactive antibody and the immediate (0-7 days) pretransplant sera. A positive donor specific IgM-AHG XM has not been a contraindication to transplant. Reports suggest that donor-specific flow cytometry cross-matches (FCXM) may be more clinically informative than the AHG-XM. METHODS We therefore evaluated the impact of a positive FCXM (IgG or IgM) on the rejection frequency (0-12 months after transplant) and 1-year graft survival for cyclosporine-prednisone-treated primary (haplo-ID and CAD) renal allograft recipients. All transplants were performed after a negative donor-specific IgG AHG-XM regardless of the IgM-AHG XM status. RESULTS Rejection frequencies (26% vs. 31%, P = NS) and 1-year graft survivals (92% vs. 89%, P = NS) were comparable for haplo-ID LRD FCXM-negative and IgG-FCXM-positive recipients. However, IgM-FCXM-positive LRD recipients experienced significantly fewer rejections (13% vs. 26% P<0.02) and an improved 1-year graft survival (100% vs. 92%, P<0.02) than FCXM-negative LRD recipients. Similar results were observed for primary CAD recipients. Rejection frequencies (40% vs. 44%, P = NS) and 1-year graft survivals (83% vs. 81%, P = NS) were comparable for primary CAD FCXM-negative and IgG-FCXM-positive recipients. Again, IgM-FCXM-positive primary CAD recipients experienced significantly fewer rejections (22% vs. 40%, P<0.02) and improved 1-year graft survivals (89% vs. 83%, P<0.05) than FCXM-negative recipients. CONCLUSION These data suggest that, after a negative donor-specific IgG-AHG XM, an IgG-positive FCXM is not a contraindication to transplantation. The presence of IgM may be beneficial in reducing the occurrence of rejection episodes and improving graft survivals.

The role of flow cytometry-detected IgG and IgM anti-donor antibodies in cardiac allograft recipients.

BACKGROUND At our transplant center, cardiac allograft recipients undergo transplantation following a negative IgG anti-human globulin (AHG) crossmatch (XM). Flow cytometry crossmatching (FCXM) is a more sensitive XM procedure than the AHG XM procedure, yet there is limited information regarding the clinical relevance of FCXM to cardiac allograft outcome. METHODS FCXM was performed retrospectively using the pretransplant sera from 140 recipients of primary cardiac allografts who underwent transplantation after AHG-IgG-NEG XM. The FCXM results were correlated to posttransplant rejection and patient survival. RESULTS All of the patients were auto-XM-NEG. Twenty-two of 140 patients (16%) displayed IgG(+) FCXM and had a significantly poorer 1-year survival rate than did 57 of the FCXM-NEG recipients (68% vs. 86%, P<0.02). Moreover, 50% of the IgG(+) FCXM recipients experienced early rejections (< or =14 days postoperatively) compared with only 16% for the FCXM-NEG recipients (P<0.01). The survival rate of 92% for IgM(+) FCXM recipients (n=37) was significantly improved compared with the 86% survival rate for FCXM-NEG control recipients (P<0.05), suggesting a protective role for IgM. Consistent with this interpretation is that the 1-year survival rate of 79% for the IgG, IgM FCXM(+) recipients (n=24) was significantly better than the 68% survival rate for the IgG(+) FCXM recipients (P<0.02). CONCLUSIONS These data suggest that IgG(+) FCXM identifies a subset of AHG-IgG-NEG XM cardiac allograft recipients who are at risk for early rejections and poor survival. In contrast, the presence of IgM may be beneficial to survival.

Can an immunologically, nonreactive potential allograft recipient undergo transplantation without a donor-specific crossmatch?

Performance of the pretransplant crossmatch requires 4 or more hours . Delays in the crossmatch might alter operating room availability and thereby increase donor organ cold ischemia time that might then result in increased risk of delayed graft function. To avoid these problems, recipients could be identified who would be expected to display negative donor crossmatches and who could be transplanted with a concurrent or retrospective rather than a pretransplant crossmatch. We, therefore, evaluated the percent reactive antibodies and donor IgG-antihuman globulin (AHG) crossmatch results of 1165 sera from 220 potential allograft recipients. Twenty-five (11%) of 220 recipients consistently displayed a 0% PRA and, with only one exception, their sera (n= 156) tested IgG-AHG crossmatch-negative against potential cadaveric donors (a 0.6% IgG-AHG positive crossmatch risk). These data suggest that the timing of the pretransplant serum crossmatch could be altered for a highly selected group of immunologically nonreactive recipients.

Comparison between enzyme-linked immunosorbent assay and cytotoxic cross-match procedures for detecting IgG anti-donor antibodies

BACKGROUND Disadvantages inherent to complement-dependent cytotoxicity cross-match (CDC XM) methods are the requirements for complement and viable target cells, detection of antibodies (Abs) against non-HLA antigens, and subjective scoring. Cross-Stat (SangStat Medical Corp., Menlo Park, CA), a recently developed enzyme-linked immunosorbent assay XM procedure for the detection of IgG anti-donor HLA Abs, is theoretically devoid of these flaws. METHODS We compared results of Cross-Stat and our standard anti-human globulin (AHG)-enhanced CDC XM procedure on 524 sera from 230 transplant candidates, which were evaluated against 51 cadaveric donors. RESULTS There was a significant correlation between AHG-CDC IgG XM and Cross-Stat results (P<0.001). For false negative sera, repeat AHG-CDC IgG XMs were still positive after platelet absorption, indicating that the Abs present were either non-HLA Abs or anti-HLA class II. Flow cytometry testing of false positive sera usually (42/62) substantiated Cross-Stat results, indicating that the discrepancy with AHG-CDC IgG XM is caused by greater sensitivity of Cross-Stat. Relative to the AHG-CDC XM, the sensitivity of Cross-Stat was 100%, the specificity was 93%, the positive predictive value was 73%, and the negative predictive value was 100%. A technical shortcoming of the Cross-Stat assay is that the frequency of indeterminate samples in the assays was 15%. Among 49 Cross-Stat negative vs. 13 Cross-Stat positive primary cadaveric renal allograft recipients (all AHG-CDC IgG-XM negative), there was no statistical difference in overall graft survival. CONCLUSION Given the important theoretical advantages of enzyme-linked immunosorbent assay-based XM methods over the CDC XM, however, further testing of the clinical relevance of the Cross-Stat is warranted.

Implications of ABO error rates in proficiency testing for solid organ transplantation

ABO poses an important histocompatibility barrier in solid organ transplantation. The classical ABO blood group barrier is defined by isoagglutinins that are naturally present in A, B, and O patients. The antibodies are directed respectively, to the blood group antigens, B, A, and A and B, which are located on the vascular endothelium of the transplanted organ. Without therapy to lower the isoagglutinin levels, ABO incompatible transplantation usually results in hyperacute or accelerated acute rejection of the organ (1–3) as the result of antibody binding to the blood group antigen on the graft vascular endothelium, complement fixation, and, ultimately, graft thrombosis. Such an event was brought to national attention by the media in 2003 when a patient died due to complications of receiving an ABO-incompatible heart/lung transplant. The United Network for Organ Sharing (UNOS) responded by mandating that when any patient was added to the waiting list they must be ABO typed two times and two people must separately enter the patient’s results into the UNOS computer (UNOS Policy 3.1.4.2, www. unos.org). UNOS also mandated that deceased donors from whom organs are transplanted must be typed twice and their blood type entered into the UNOS system by two individuals (UNOS Policy 5.0). Those changes were warranted, but at and since that time, no ABO discrepancy data have been published so the relative risk of ABO typing errors cannot be assessed. The true incidence of errors in ABO typing of transplant donors and recipients is not known. We know that errors do occur throughout the complex process of transplantation, which have resulted in ABO-incompatible transplantation (1–3). Due to the many points in that process’ error continuum, such uncertainties due to under-reporting, undetected cases in which an error occurred without ABO incompatibility, an ABOincompatibility occurred but was not recognized as such, verbal mis-communication of ABO results, among others, the true frequency of errors is practically impossible to measure. The primary aim of this study was to examine the ABO typing error rate of the laboratories’ analytical and postanalytical phases of ABO typing. We did that by evaluating the ABO discrepancy data from the College of American Pathologists (CAP) HLAS and J Transfusion medicine proficiency testing surveys. If performed in accord with CAP standards, these survey data should closely approximate the actual ABO error rate, or discrepancy rate for routine laboratory ABO testing for solid organ transplantation, since this is a large collection of ABO proficiency testing data ( 116,000 tests) that were blindly tested by more than 3,000 labs over a four year time period (2000 to 2004). The second aim was to evaluate discrepancy rates for A subgrouping (A1 and A2) since more centers are transplanting kidneys and other organs from blood group A2 and A2B donors into blood group B patients (3–5) who wait longer for a kidney than any other blood group (6).

Retransplantation following AHG-negative crossmatches

Retransplant patients are an immunologically unique group since they have already experienced graft loss and have been exposed to a bolus of alloantigen. Having previously lost a transplant, these patients are often sensitized and have poorer early graft function resulting in patient management problems [1, 2]. Survival rates for cadaveric retransplantations have been consistently lower than those for primary grafts [3, 4, 5, 6]. Some of the risk factors associated. with poor retransplantation outcome include patient high PRA, recipient-donor HLA mismatches, positive flow cytometry crossmatches and, perhaps the most important, previous graft survival time less than 3–6 months [3, 4, [7, 8, 9]. Since the introduction of cyclosporine based immunosuppressive protocols in 1983, the one year regraft survival rate of second cadaver donor renal allografts has remained relatively constant, whereas that of first transplants has significantly improved [9]. However, since 1988 survival rates for retransplant cadaver-donor grafts have improved each year with the difference between first and second transplant one year survival rates decreasing from 8% in 1988 to 2% in 1991 [10]. These recent improvements may reflect the impact of more efficacious immunosuppression and more sensitive and informative crossmatching [6, 11]. However, data from the UNOS Registry suggests that a decreasing number of high-risk patients, especially reTx patients with short previous graft survival times, have been retransplanted in recent years and that selection of more low-risk reTx candidates may be an important contributing factor in explaining improved regraft survivals [8].