Positive Complement-dependent Cytotoxicity Crossmatch Research Articles

Complement-dependent cytotoxicity crossmatch in solid organ transplantation: The gold standard or golden history?

Complement-dependent cytotoxicity crossmatch (CDC-XM) has been considered for many years the standard of practice for determining compatibility in solid organ transplantation (SOT). However, as this method is laborious, time intensive and lacks sensitivity and specificity, it has been replaced in many laboratories worldwide by flow cytometry crossmatch (FCXM) and/or virtual crossmatch (vXM).With this study we intend to show the relevance of performing CDC-XM in the era of virtual crossmatching. We retrospectively analyzed 1,007 consecutive T and B cell deceased donor (DD) CDC-XMs performed in parallel using non-treated and dithiothreitol (DTT) treated sera between May 2022 and January 2023 in waitlisted patients with no donor specific antibodies (DSA) against HLA-A, B and/or DR antigens. Thirty five of 1,007 (3.5%) T cell crossmatches and 132 of 1,007 (13.1%) B cell crossmatches were positive with non-treated sera. Correlation with the vXM demonstrated no DSA in any of the positive T cell crossmatches. DSA were also absent in 126/132 positive B cell crossmatches, indicating a high rate of false positive CDC-XM. Indeed, only 4/35 T cell and 13/132 B cell CDC-XM remained positive after treatment with DTT, confirming that false positive reactivity with non-treated sera is high. Class I HLA DSA against C locus antigens were present in 17/1,007 T cell crossmatches and none were detected by CDC-XM (sensitivity = 0%). Similarly, only 6/77 B cell crossmatches with DSA targeting HLA-C, DQ and/or DP antigens were CDC-XM positive (sensitivity = 7.8%). Furthermore, only 4/6 positive B cell CDC-XM were confirmed to have complement binding potential using the C1q assay, suggesting additional false positive reactivity in 2/6 of the positive CDC-XM.Our study demonstrates that CDC-XM exhibits poor sensitivity, high false positive reactivity (especially without DTT treatment) and does not meaningfully contribute to pre-transplant compatibility testing in the context of vXM based allocation. Furthermore, the use of CDC-XM can unnecessarily delay or even prevent safe and appropriate transplant allocation.

Impact of allele-specific anti–human leukocyte antigen class I antibodies on organ allocation

Technological advances in the field of histocompatibility have allowed us to define anti–human leukocyte antigen (HLA) antibody specificity at the allelic level. However, how allele-specific antibodies affect organ allocation is poorly studied. We examined allelic specificities of class I HLA antibodies in 6726 consecutive serum samples from 2953 transplant candidates and evaluated their impact on the corresponding crossmatch and organ allocation. Out of 17 class I HLA antigens represented by >1 allele in the LABScreen single antigen bead assay, 12 had potential allele-specific reactivity. Taking advantage of our unbiased cohort of deceased donor-candidate testing (123,135 complement-dependent cytotoxicity crossmatches between 2014 and 2017), we estimated that the presence of allele-specific antibody detected using a single antigen bead assay (median fluorescence intensity, >3000) against only the rare allele was a poor predictor of a positive complement-dependent cytotoxicity crossmatch, with a positive predictive value of 0% to 7%, compared with 52.5% in allele-concordant class I HLA antibodies against A or B locus antigens. Further, we confirmed allele-specific reactivity using flow crossmatch in 3 scenarios: A11:01/A11:02, A68:01/A68:02, and B44:02/B44:03. Our results suggest that allele-specific antibodies may unnecessarily exclude transplant candidates (up to 10%) from organ offers by overcalling unacceptable antigens; incorporation of selective reactivity pattern in allocation may promote precision matching and more equitable allocation.

Correlation of Luminex-Based Single Antigen Based Results With Complement-Dependent Cytotoxicity Crossmatch and Flow Cytometry Crossmatch Results: A Single-Center Experience From Istanbul

This study aimed to retrospectively investigate the correlation of mean Class I donor-specific antibody (DSA) intensity values detected in Luminex-based techniques with the results of complement-dependent cytotoxicity crossmatch (CDC-XM) and flow cytometry crossmatch (FC-XM) results. A total of 335 patients with kidney failure and their living donors whose CDC-XM, FC-XM, and single antigen based (SAB) tests were studied between 2018 and 2020 for transplant preparation from living donor candidates were included in the study. Patients were divided into 4 groups according to their mean fluorescence intensity (MFI) values of SAB assay. Anti-HLA antibodies (class I and/or class II) were detected using SAB in 91.6% patients included in the study (MFI >1000). Class I DSA was positive in 34.8% of patients with anti-HLA antibodies. When CDC-XM and FC-XM results were evaluated in the 4 groups separated according to MFI values, 3 patients with DSA MFI <1000 had negative CDC-XM and T-B-FC-XM results. Of 32 patients with DSA-MFI between 1000 and 3000, 93.75% (n=30) had T-B-FC-XM or CDC-XM-negative results, and 6.25% (n=2) had B-FC-XM-positive results. The CDC-XM, T, and B-FC-XM were negative in all 17 patients with DSA-MFI between 3000 and 5000. Our results showed that MFI >5834 DSA values were significantly correlated with positive T-FC-XM (P < .001), and MFI >6016 values were significantly correlated with positive CDC-XM (P=.002). In addition, MFI values >5000 were associated with both CDC-XM and FC-XM in our study. The MFI values >5000 correlated with both CDC-XM and FC-XM.

Desensitization using IVIG alone for living-donor kidney transplant: impact on donor-specific antibodies.

Sensitization to human leukocyte antigen is a barrier to. Few data have been published on desensitization using polyvalent human intravenous immunoglobulin (IVIG) alone. We retrospectively reviewed the of 45 patients with a positive complement-dependent cytotoxicity crossmatch (CDCXM) or flow cytometry crossmatch (FCXM) against living donors from January 2003 to December 2014. Of these, 12 were excluded. Patients received monthly IVIG infusions (2 g/kg) only until they had a negative T-cell and B-cell FCXM. During the 33 patients, 22 (66.7%) underwent living donor kidney transplantation, 7 (21.2%) received a deceased donor graft, and 4 (12.1%) did not undergo transplantation. The median class I and II panel reactive antibodies for these patients were 80.5% (range 61%-95%) and 83.0% (range 42%-94%), respectively. Patients (81.8%) had a positive T-cell and/or B-cell CDCXM and 4 (18.2%) had a positive T-cell and/or B-cell FCXM. Patients underwent transplantation after a median of 6 (range 3-16). The median donor-specific antibody mean fluorescence intensity sum was 5057 (range 2246-11,691) before and 1389 (range 934-2492) after desensitization (p = 0.0001). Mean patient follow-up time after transplantation was 60.5 (SD, 36.8) months. Nine patients (45.0%). Death-censored graft survival at 1, 3, and 5 years after transplant was 86.4, 86.4, and 79.2%, respectively and patient survival was 95.5, 95.5, and 83.7%, respectively. Desensitization using IVIG alone is an effective strategy, allowing successful transplantation in 87.9% of these highly sensitized patients.

Association of Panel Reactive Antibodies (PRA) with Complement Dependent Cytotoxicity (CDC) Cross-Match in Pre-Renal Transplant Recipients

Objective: To determine the association of panel reactive antibodies (PRA) with complement-dependent cytotoxicity (CDC) cross-match in Pakistani pre-renal transplant recipients.&#x0D; Study Design: Cross-sectional study.&#x0D; Place and Duration of Study: Immunology Department, Armed Forces Institute of Pathology, Rawalpindi from Oct 2017 to Oct 2018.&#x0D; Methodology: A total of 162 patients referred to the Department of Immunology for pre-transplant workup for renal transplantation were included. Informed consent was taken, and detailed history was recorded. Frequency and percentages were calculated for cross-match positivity and most frequent anti-HLA antibodies.&#x0D; Results: Panel reactive antibodies (PRA) were present in 48 patients (30%), while complement-dependent cytotoxicity (CDC) cross-match was positive in 16 patients (10%). Out of 141 male patients, 35 (25%) were positive for PRA, while 10 (7%) had positive CDC cross-match. Out of 21 female recipients, 13 (62%) were positive for PRA, and 6 (28%) had positive CDC crossmatch. One male patient positive for CDC cross-match was negative for PRA. Patients positive for both CDC cross-match and PRA have an average mean fluorescent intensity (MFI) of more than 4000. CDC cross-match and PRA were strongly associated, whereas no significant association was found between CDC cross-match and anti-MIC antibodies.&#x0D; Conclusion: Complement dependent cytotoxicity (CDC) cross-match and panel reactive antibodies (PRA) should be routine in patients undergoing renal transplants as alone CDC cross-match can give false negative or false-positive results. At the same time, CDC cross-match lacks detection of anti MIC antibodies involved in graft rejection.

The utility of cytolytic flow cytometry crossmatch before kidney transplantation

The donor/recipient matching in kidney transplantation is based on approved laboratory tests, which are complement-dependent cytotoxic crossmatch (CDC-XM) and flow cytometry crossmatch (FCXM). Both have some disadvantages: CDC-XM has low sensitivity, whereas FCXM does not differentiate between lytic vs. non-lytic alloantibodies. To find an improved method, we have developed a new crossmatch technique of cytolytic flow cytometry crossmatch (cFCXM), which allows for sensitive detection of clinically relevant complement-binding antibodies. The cFCXM assay detects dead cells with viability dye that ensue from the binding of allospecific lytic antibodies. In our study, 135 unsensitized kidney transplant recipients were recruited based on the CDC-XM and FCXM results and the clinical utility of cFCXM was evaluated. The 5-year follow-up for acute rejection incidents revealed that cFCXM could verify the clinical relevance of positive FCXM results as recipients with positive FCXM but negative CDC-XM had the same risk of rejection as patients with both negative CDC-XM/FCXM results. These findings suggest that cFCXM assay may provide more precise immunological risk assessment in kidney transplant recipients.

Presence of a survival benefit of HLA-incompatible living donor kidney transplantation compared to waiting or HLA-compatible deceased donor kidney transplantation with a long waiting time

HLA-incompatible living donor kidney transplantation (LDKT) is one of efforts to increase kidney transplantation opportunity for sensitized patients with kidney failure. However, there are conflicting reports for outcomes of HLA-incompatible kidney transplantation compared to patients who wait for HLA-compatible deceased donor kidney transplantation (DDKT) in the United States and United Kingdom. Waiting for an HLA-compatible DDKT is relatively disadvantageous in Korea, because the average waiting time is more than five years. To study this further, we compared outcomes of HLA-incompatible LDKT with those who wait for HLA-compatible DDKT in Korea. One hundred eighty nine patients underwent HLA-incompatible LDKT after desensitization between 2006 and 2018 in two Korean hospitals (42 with a positive complement-dependent cytotoxicity cross-match, 89 with a positive flow cytometric cross-match, and 58 with a positive donor-specific antibody with negative cross-match). The distribution of matched variables was comparable between the HLA-incompatible LDKT group and the matched control groups (waiting-list-only group; and the waiting-list-or-HLA-compatible-DDKT groups; 930 patients each). The HLA-incompatible LDKT group showed a significantly better patient survival rate compared to the waiting-list-only group and the waiting-list-or-HLA-compatible-DDKT groups. Furthermore, the HLA-incompatible LDKT group showed a significant survival benefit as compared with the matched groups at all strength of donor-specific antibodies. Thus, HLA-incompatible LDKT could have a survival benefit as compared with patients who were waitlisted for HLA-compatible DDKT or received HLA-compatible DDKT in Korea. This suggests that HLA-incompatible LDKT as a good option for sensitized patients with kidney failure in countries with prolonged waiting times for DDKT.

THU270 - Positive complement-dependent cytotoxicity crossmatch results do not influence long-term graft or patient survival following liver transplantation

THU270 - Positive complement-dependent cytotoxicity crossmatch results do not influence long-term graft or patient survival following liver transplantation

Clinical Prognosis of Renal Retransplant Patients: A Single-Center Experience

BackgroundRetransplantation is a treatment option in patients with end-stage renal failure due to graft loss. Outcomes of these patients due to high immunologic risk remain unclear. The aim of this study was to evaluate outcomes of renal retransplantation patients retrospectively. MethodsRenal retransplant patients in our unit were evaluated retrospectively between 2010 and 2018. Patients’ demographic characteristics, primary diseases, the causes of prior graft loss, immunologic status, desensitization protocols, the induction and maintenance treatments, the complications during the follow-up period, numbers of acute rejections, and the clinical prognosis were all detected from the patients’ files. ResultsWe retrospectively evaluated 17 patients who underwent a second or third renal allograft. Of these, 16 received a second and the remaining 1 patient received a third renal allograft. Immunologically, all of the 17 patients had negative flow cytometry crossmatch, 1 patient had a positive complement-dependent cytotoxicity crossmatch (Auto 12%), 16 patients had positive panel reactive antibody, the median HLA-mismatch was 3.5, and the score of donor-specific antibody relative intensity score (RIS) was 6.4 ± 6.3. Ten pretransplant patients had desensitization treatment. While scores for HLA-MM and HLA-RIS in the patients who had a desensitization therapy were determined higher, no statistical difference was observed (respectively, P = .28 and .55). No acute rejection episode developed. BK virus DNA viremia was detected in 4 patients during the posttransplant 6th month. We observed no patient death or no graft loss during the follow-up period. ConclusionAlthough the retransplant patients who had a graft loss previously have high immunologic risks, retransplantation is reliable in these patients, but they should be followed up carefully in terms of BKV nephropathy.

Successful renal transplantation in a patient with perinuclear antineutrophil cytoplasmic antibody-associated vasculitis with chronic kidney disease with complement-dependent cytotoxicity crossmatch positivity (autoantibody induced) and donor-specific antibodies and flow cytometry crossmatch negative

The positive complement-dependent cytotoxicity crossmatch (CDCXM) is considered as a contraindication for renal transplantation (RT) since long in an effort to avoid immediate rejection. There has been tremendous development in our understanding of transplant immunology today, and more sensitive and specific methods such as flow cytometry crossmatch (FCXM) and solid-phase antibody screening are used for detailed immunological assessment. We report successful RT in a 23-year-old girl with end-stage renal disease due to vasculitis on dialysis for 2 years. Before transplantation, the patient had Anti Human Globulin CDCXM positive while Dithioerythritol (DTT)-CDCXM was negative. The patient had no donor-specific antibodies examined by Luminex single-antigen beads and her FCMXM was negative. In the posttransplant period, there was no evidence of immune injury. Her serum creatinine was 0.7 mg/dl on the 3rd posttransplant day at the time of discharge. Induction immunosuppression was rabbit thymoglobulin (1.5 mb/kg) and methylprednisolone (500 mg, 3 doses) and maintenance immunosuppression was tacrolimus + prednisolone + mycophenolic acid. We found that even though CDCXM positivity has been traditionally considered as a contraindication for renal transplantation, in few carefully selected patients, we can still proceed to renal transplantation even if CDCXM is positive if more advanced and robust immunological tools such as FCMXM and donor-specific antibodies are negative. In our patient, AHG-CDCXM was positive while DTT-CDCXM was negative making it clear that antibodies of IgM origin were probably responsible for CDCXM positivity.

Comparative analysis of Luminex-based donor-specific antibody mean fluorescence intensity values with complement-dependent cytotoxicity & flow crossmatch results in live donor renal transplantation.

Background & objectives:Antibodies specific to donor human leucocyte antigen (HLA) play a critical role in graft rejection and graft loss. In recent years, techniques for their detection have evolved significantly providing an ever-increasing degree of sensitivity and specificity, from the conventional cell-based assays to the advanced solid-phase system based on the Luminex platform. Consensus is still evolving on the routine employment of all these methods, either stand alone or in combination. The objective of this study was to explore the near-accurate mean fluorescence intensity (MFI) cut-off values detected on Luminex platform predicting the strength of cell-based crossmatch results.Methods:Serum samples from 116 primary renal transplant recipients awaiting transplantation were tested for the presence of antidonor antibodies by the complement-dependent cytotoxicity (CDC) and flow crossmatch (FCXM) methods with their corresponding donors as well as for HLA-donor-specific antibodies (DSA) detection using a sensitive single antigen bead (SAB) assay.Results:None of the patients having HLA Class I DSA with MFI values <1000 showed positivity for T-cell FCXM or CDC crossmatch, while in the group having MFI values between 1000 and 3000, 54 per cent showed positivity for the FCXM but none by the CDC method. However, in the group having MFI values >3000, 95 per cent of cases were positive for FCXM. Further, those groups with MFI values between 3000 and 5000, only 36 per cent were positive for CDC crossmatch, while 90 per cent showed positivity in the group with MFI >7000.Interpretation & conclusions:A cut-off MFI value of 3000 for Luminex SAB-based assay was found to significantly correlate with the FCXM positivity while a MFI value of 7000 and above predicted a positive CDC crossmatch. MFI cut-off value obtained as a surrogate marker for CDC and FCXM tests will help in resolving the limitations of different cell-based techniques.

Case Report: Anti-HLA-DQα Antibodies Have Clinical Impacts in Highly Immunized Kidney Patients

Context and purpose: Organ exchange programmes are important in order to ensure organs to highly immunized kidney patients. In this case report, we describe a highly immunized patient in an acceptable mismatch program who had an unexpected positive Complement-Dependent Cytotoxicity (CDC) crossmatch with a deceased donor exchange.Main findings: Subsequent evaluation revealed donor specific antibodies characterized by serological epitopes, which included the HLA-DQα chain, were masked in the standard IgG Luminex assay unless the serum was diluted; however, the antibodies demonstrated high MFI values when the serum was pretreated with EDTA or analyzed with the C1q assay. These findings led us to evaluate the extent of masked anti-HLA-DQ antibodies that included HLADQα/ DQβ pairs rather than just the HLA-DQβ chains in 16 highly immunized patients on the kidney transplantation waiting list at our transplantation center. We determined that 25% of the patients had antibodies directed towards serological epitopes that involved the anti-HLA-DQα chain but were masked in the standard IgG assay.Principal conclusions: We conclude that masked anti-HLA-DQ antibodies directed towards serological epitopes that also involves the HLA-DQα chain are common and should be considered as they may cause a positive CDC crossmatch. Thus, donor specific HLA-DQ antibodies must be evaluated in relation to full HLA-DQα; DQβ pairs in the serological molecules of the donor.

P152 Prozone phenomena may skew the strength of Donor Specific Antibody (DSA)

Aim Positive Complement Dependent Cytotoxicity (CDC) and Flow Cytometry Crossmatch (FCXM) usually correlate with high titer of Donor Specific Antibodies (DSA). Now a days assignment of relevant DSA is based on solid phase assay (SPA), in which false low/negative results can be observed due to prozone effect. Here we present three cases underwent a workup for living related kidney transplant that their crossmatch data does not correlate with the single antigen (SA) Median Fluorescent Intensity (MFI) value. Method B cell and/or T cell AHG-CDC, FCXM and antibody identification by SPA were performed for three living related pairs. Results Two patients show positive T cell AHG-CDC crossmatch and one shows positive B cell CDC crossmatch with 8 reaction strength at 1:8 dilution and positive FCXM with very high Median Channel Shift (MCS). However the DSA result was negative for one patient and the other two patients showed A2 (891 MFI) and DQ6 (1299 MFI) respectively. The SA testing indicated possible skewed MFI value, therefore all serum were diluted 1:4 to overcome a prozone effect if present. Conclusion MFI value of the SPA was not correlated with the high MCS and positive CDC crossmatch in undiluted serum due prozone phenomena of high antibody concentration. Therefore serum dilution should be considered if strong crossmatch reactivity observed.

Deceased donor kidney transplantation across donor-specific antibody barriers: predictors of antibody-mediated rejection

Apheresis-based desensitization allows for successful transplantation across major immunological barriers. For donor-specific antibody (DSA)- and/or crossmatch-positive transplantation, however, it has been shown that even intense immunomodulation may not completely prevent antibody-mediated rejection (ABMR). In this study, we evaluated transplant outcomes in 101 DSA+ deceased donor kidney transplant recipients (transplantation between 2009 and 2013; median follow-up: 24 months) who were subjected to immunoadsorption (IA)-based desensitization. Treatment included a single pre-transplant IA session, followed by anti-lymphocyte antibody and serial post-transplant IA. In 27 cases, a positive complement-dependent cytotoxicity crossmatch (CDCXM) was rendered negative immediately before transplantation. Seventy-four of the DSA+ recipients had a negative CDCXM already before IA. Three-year death-censored graft survival in DSA+ patients was significantly worse than in 513 DSA- recipients transplanted during the same period (79 versus 88%, P = 0.008). Thirty-three DSA+ recipients (33%) had ABMR. While a positive baseline CDCXM showed only a trend towards higher ABMR rates (41 versus 30% in CDCXM- recipients, P = 0.2), DSA mean fluorescence intensity (MFI) in single bead assays significantly associated with rejection, showing 20 versus 71% ABMR rates at <5000 versus >15 000 peak DSA MFI. The predictive value of MFI was moderate, with the highest accuracy at a median of 13 300 MFI (after cross-validation: 0.72). Other baseline variables, including CDC assay results, human leukocyte antigen mismatch, prior transplantation or type of induction treatment, did not add independent predictive information. IA-based desensitization failed to prevent ABMR in a considerable number of DSA+ recipients. Assessing DSA MFI may help stratify risk of rejection, supporting its use as a guide to organ allocation and individualized treatment.

Desensitization Before Living Donor Kidney Transplantation in Highly HLA-Sensitized Patients: A Single-Center Study

BackgroundRenal transplantation in highly sensitized patients represents a major clinical challenge leading to long periods on the waiting list. When a living donor is available, the use of different strategies to desensitize recipients with preformed human leukocyte antigen antibodies can allow a successful transplantation. MethodsWe performed a retrospective observational study including all living donor kidney transplantation (LDKT) with desensitization (DS) from 2008 to 2014 in our transplant unit. The rates of rejection and graft survival were evaluated. DS consisted of plasma exchange (PE), rituximab (RTX), and intravenous immunoglobulin (IVIG) induction with thymoglobulin and maintenance immunosuppression with tacrolimus, corticosteroids, and mycophenolate mofetil. ResultsFrom 2008 to 2014, we performed 368 LDKT, with 31 receiving desensitization. Seven cases from a clinical trial were excluded. Demographic data and outcomes were recorded. All of the patients received RTX + PE + IVIG. DS was performed for positive complement-dependent cytotoxicity cross-match (4.2%), T-cell– and/or B-cell–positive flow cytometry cross-match (87.5%) and presence of donor-specific antibodies alone (8.3%). We identified 23 episodes of rejection in 12 patients (50%); 79% were antibody-mediated rejections (AMR). Graft failure was 12.5%, with a mean time to graft loss of 229 ± 203 days. Mean follow-up was 37 ± 27 months, and graft survival was 91% and 86% at 1 and 5 years, respectively. ConclusionsDesensitization in LDKT appears to offer an acceptable option for highly sensitized patients. In our series, 41% presented an AMR and 12.5% showed transplant glomerulopathy in protocol and/or indication biopsies. However, short-term outcomes and graft survival were satisfactory.

P114 : CHANGE OF THE PATIENT’S ANTI HLA ANTIBODY PROFILE BETWEEN INITIAL AND FINAL WORK-UP

Aim Patients anti HLA antibody profile could change overtime in its specificity, strength and/or behavior. Here we are presenting a case for a patient with end stage renal disease (ESRD) workup for living related kidney transplant, in which antibody profile changed between the initial and final work-up (the day before transplant). Methods HLA Class I highly sensitized patient with anti A26 and anti B50 Donor Specific Antibody (DSA) with 2904, 12168 Median fluorescent Intensity (MFI) respectively, underwent desensitization protocol with Rituximab, high dose IVIG and Plasma Apheresis. One day before transplantation, final Complement Dependent Cytotoxicity crossmatch (CDCXM) as well as Flow Cytometry Crossmatch (FCXM) were performed utilizing historical serum “initial work-up sera prior to desensitization” and current serum post desensitization. Results As can be seen in the table below, the characteristics of the patients’ DSA changed between the initial and final work-up. The DSA changed from complement fixing antibody with a positive CDCXM crossmatch to non-complement fixing antibodies with negative CDCXM crossmatch. Conclusion This case highlights the relevant of frequent testing of anti-HLA antibodies in renal transplant candidates. As seen above, the antibody Isotype switched from fixing to none complement fixing. Thereafter, such conversion could modify the treatment modality. Result. Initial work-up Final work-up CDCXM Positive Negative FCXM Positive Positive Undiluted sera A26 (2904 MFI), B50 (12168 MFI) A26 (12421 MFI), B50 (984 MFI) Diluted sera (1:4) A26 (16680 MFI), B50 (6247 MFI) A26 (5936 MFI), B50 (Negative) C1q Positive Negative

Long-term outcomes of kidney transplantation across a positive complement-dependent cytotoxicity crossmatch.

More than 30% of potential kidney transplant recipients have pre-existing anti-human leukocyte antigen antibodies. This subgroup has significantly lower transplant rates and increased mortality. Desensitization has become an important tool to overcome this immunological barrier. However, limited data is available regarding long-term outcomes, in particular for the highest risk group with a positive complement-dependent cytotoxicity crossmatch (CDC XM) before desensitization. Between 2002 and 2010, 39 patients underwent living-kidney transplantation across a positive CDC XM against their donors at our center. The desensitization protocol involved pretransplant immunosuppression, plasmapheresis, and low-dose intravenous immunoglobulin±rituximab. Measured outcomes included patient survival, graft survival, renal function, rates of rejection, infection, and malignancy. The mean and median follow-up was 5.2 years. Patient survival was 95% at 1 year, 95% at 3 years, and 86% at 5 years. Death-censored graft survival was 94% at 1 year, 88% at 3 years, and 84% at 5 years. Uncensored graft survival was 87% at 1 year, 79% at 3 years, and 72% at 5 years. Twenty-four subjects (61%) developed acute antibody-mediated rejection of the allograft and one patient lost her graft because of hyperacute rejection. Infectious complications included pneumonia (17%), BK nephropathy (10%), and CMV disease (5%). Skin cancer was the most prevalent malignancy in 10% of patients. There were no cases of lymphoproliferative disorder. Mean serum creatinine was 1.7±1 mg/dL in functioning grafts at 5 years after transplantation. Despite high rates of early rejection, desensitization in living-kidney transplantation results in acceptable 5-year patient and graft survival rates.

Systemic Lupus Erythematosus Leading to Terminal Renal Failure and Excluding Patients from Kidney Allocation Due to Inadequate CDC-based Cross-Matching: Is There a Diagnostic Way out?

Systemic lupus erythematosus (SLE) is known to proceed to clinically relevant nephritis in more than 50% of patients and, in about 20% of these patients, to terminal renal failure. Thus, renal replacement therapy including kidney allografting is required for a considerable number of SLE-patients. For allografting patients’ donor-specific antibodies against HLA molecules of given donors (DSA) have to be excluded as preformed antibodies against these molecules represent the main cause for hyper-acute or acute rejections. In order to select recipients without these deleterious antibodies the complement-dependent cytotoxicity crossmatch (CDC-XM) assay was developed about forty years ago. Its negative pre-transplant outcome is currently regarded as the most important requirement for a successful kidney graft survival. During the last years several disadvantages of the CDC-based procedure have increasingly been discussed with respect to this assay’s high susceptibility to disruptive factors mainly leading to false positive outcomes. As is clearly shown by our case series this holds also true for the underlying disease SLE. We here present the data of SLE-patients initially destined for cadaver kidney donations. They all exhibited positive CDC-XM outcomes for the most part without known historical immunizing events. Furthermore, solid phase-based antibody screening and specification analyses did generally not show anti-HLA antibodies or antibodies directed against HLA-phenotypes of the corresponding donors thus leading to negative results of the so-called virtual crossmatch. Since exclusive virtual cross-matching is not allowed for the acceptance of allografting all of the positive CDC-XM assays were performed as reruns using alternative solid phase- (ELISA-) based crossmatch assays. In best accordance with virtual cross-matching solid phase-based cross-matching did not exhibit DSA. Our data clearly demonstrate the benefit of alternative ELISA-based cross-matching to circumvent CDC-based artefacts and point on the urgent requirement to substitute the historical CDC-based procedure at least for this group of patients.

21-P

Aim The aim of this study was to establish whether the presence of DSA detected by Luminex based solid-phase assay (SPA) correlates with the cross-match results performed by complement-dependent cytotoxicity (CDC) in deceased donor kidney transplantation. Methods Between January 1, 2012 and December 31, 2012, lymphocytes from a total of 398 deceased donors were cross-matched by our laboratory. For each donor, sera from an average of 6.24 patients were selected for direct cross-matching by CDC, amounting to a total of 2,485 cross-matches. Pre-transplant sera were screened by CDC on HLA reference panel, and by LABScreen using HLA class I and class II antigen coated beads. Results Of the 2,485 patients cross-matched, 870 had DSA and should have been excluded as potential transplant recipients on this basis. However, 39.3% (342/870) of the DSA positive population showed no cytotoxic antibodies to the putative donor reflecting the non-Complement fixing nature of the antibodies detected by SPA and/or their low affinity for their target antigen. Since 60.7% (528/870) of patients with DSA showed positive CDC cross-matches, there was still a highly significant correlation (P Conclusions In this study, we have shown that 39.3% of patients with DSA would have been denied the transplant if it was solely based on the virtual cross-match result. Furthermore, there was no statistically significant difference between graft function at one year in patients with or without DSA.

Comparison of Complement-dependent Cytotoxic and Flow-cytometry Crossmatch Results Before Cadaveric Kidney Transplantation

AimThe presence of HLA donor-specific antibodies (DSA) before kidney transplantation decreases graft survival. In this study, we compared crossmatch results of kidney transplantation candidates, for cadaveric renal donation between March 10, 2012, and September 7, 2012. Material and MethodThe 47 kidney transplantation candidates tested for crossmatch included 10 for cadaveric donor organs. Two crossmatch methods were performed: complement-dependent cytotoxic crossmatch (CDCXM) and flow cytometry crossmatch (FCXM). Spleen cells were used as the source of lymphocytes for all crossmatch tests. ResultsThe T and B cell ratios isolated from spleen were 38.8% and 34.8%, respectively. The concordance ratio of the two methods was 76.6% with 23.4% discordant results. Regarding the discordant results, 4.2% were positive CDCXM but negative FCXM; 191%, negative CDCXM but positive FCXM. All patients displaying positive crossmatches had a previous immunization history. As a result, we speculated that the positive CDCXM but negative FCXM results were due to the washing procedures in the FCXM disturbing antigen-antibody complexes. We suggest at least two different methods to be performed for crossmatch tests before kidney transplantation. CDCXM detects immunoglobulin G1 (IgG1) and IgG3, which are critical for rejection. FCXM is able to detect all IgG subgroups because of its high sensivity. As a result we suggest that both CDCXM and FCXM are preferrable strategies to detect DSAs.