Comparative Assessment of Anti-HLA Antibodies Using Two Commercially Available Luminex-Based Assays

Clerkin, Kevin J.; See, Sarah B.; Farr, Maryjane; Restaino, Susan; Serban, George; Latif, F.; Li, Lingzhi; Colombo, P.C.; Vlad, George; Ray, Bryan; Vasilescu, Elena R.; Zorn, Emmanuel

doi:10.1097/txd.0000000000000734

Cited by 20 publications

(23 citation statements)

References 17 publications

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“…In all previous studies investigating the presence of IgA HLA antibodies by luminex, One Lambda kits were used, whereas we used Lifecodes, Immucor kits. As known from the IgG HLA analyses, the two vendors differ in sensitivity, specificity, antigen panel composition, and level of denatured antigens 29‐32 . In the present study, the IgA‐LMX and IgA‐SAB results showed sometimes a discrepant reactivity, what is in line with what was previously reported for the detection of IgG HLA antibodies 33 …”

Section: Discussionsupporting

confidence: 91%

“…As known from the IgG HLA analyses, the two vendors differ in sensitivity, specificity, antigen panel composition, and level of denatured antigens. [29][30][31][32] In the present study, the IgA-LMX and IgA-SAB results showed sometimes a discrepant reactivity, what is in line with what was previously reported for the detection of IgG HLA antibodies. 33 The low incidence of IgA HLA antibodies may be caused by the competition for the binding sites on HLA molecules, existing between IgG and IgA isotype of HLA antibodies, since IgG serum concentrations are significantly higher than those of IgA.…”

Section: Discussionsupporting

confidence: 90%

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Low incidence of IgA isotype of HLA antibodies in alloantigen exposed individuals

Car

Karahan

Dreyer

et al. 2020

HLA

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Human leukocyte antigen (HLA) antibodies are induced by pregnancy, transfusion, or transplantation. Serum from transplant recipients is regularly screened for IgG HLA antibodies because of their clinical relevance for transplant outcome. While other isotypes of HLA antibodies, such as IgA may also contribute to the alloimmune response, validated detection assays for IgA HLA antibody detection are lacking. Therefore, we modified the commonly used luminex screening assay for IgG HLA antibody detection (IgG‐LMX) into an IgA HLA antibody screening assay (IgA‐LMX). Optimization and validation was performed with IgG, IgA1, and IgA2 isotype variants of HLA‐specific human recombinant monoclonal antibodies (mAbs). Reactivity patterns of IgA1 and IgA2 isotype HLA‐specific mAbs in IgA‐LMX were identical to those of the IgG isotype. Cross‐reactivity with IgG and IgM antibodies and nonspecific binding to the beads were excluded. Further assay validation showed the absence of IgA HLA antibodies in serum from individuals without alloantigen exposure (n = 18). When the IgA‐LMX assay was applied to sera from 289 individuals with known alloantigen exposure through pregnancy (n = 91) or kidney transplantation (n = 198), IgA HLA antibodies were detected in 3.5% of individuals; eight patients on the kidney retransplant waitlist and two women immunized through pregnancy. The majority (90%) of IgA HLA antibodies were directed against HLA class II and were always present in conjunction with IgG HLA antibodies. Results of this study show that this validated IgA‐LMX method can serve as a screening assay for IgA HLA antibodies and that the incidence of IgA HLA antibodies in alloantigen exposed individuals is low.

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Section: Discussionsupporting

confidence: 91%

Section: Discussionsupporting

confidence: 90%

Low incidence of IgA isotype of HLA antibodies in alloantigen exposed individuals

Car

Karahan

Dreyer

et al. 2020

HLA

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“…Furthermore, dilution of C3d+ sera showed that complement interference was not a confounder in this subanalysis. In a recent comparative study, antibody specificities detected using kits from 2 SAB vendors showed a tendency toward higher MFI values being observed on the One Lambda (OL) platform compared to Immucor . This difference may in part relate to the extra serum dilution step associated with the Immucor protocol but not in OL's, and perhaps the reason our dilution study identified only a low prevalence of complement interference in patient sera, which was already mitigated through Immucor's protocol.…”

Section: Discussionmentioning

confidence: 96%

Clinical utility of complement-dependent C3d assay in kidney recipients presenting with late allograft dysfunction

Lan

Gjertson

Zheng

et al. 2018

American Journal of Transplantation

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The objective of this study was to evaluate the utility of a complement-dependent C3d assay to risk stratify donor-specific antibodies (DSA) in a multicenter cohort of kidney recipients presenting with new-onset clinical dysfunction. A total of 106 subjects with evidence of DSA at a mean period of 5.3 ± 5.0 years posttransplant underwent testing using C3d reagents. C3d positivity was strongly associated with both the peak and sum IgG DSA MFI, with 98.3% (n = 57/58) of strongly reactive sera (peak MFI > 10 000) eliciting a positive signal. Patients with C3d+ DSA had a higher creatinine (P = .03), more significant graft fibrosis (P = .035), and a faster rate of graft loss posttest compared to those with C3d- DSA (P = .05). Subanalysis of patients with low-moderate level DSA confirmed the inferior outcome associated with C3d positivity. Despite the prognostic value of C3d as a stand-alone test, the assay did not provide independent risk prediction after incorporation of graft fibrosis in a multivariate model (P = .94). Overall, C3d offered limited discriminatory value for strong DSA with peak IgG MFI > 10 000 and in patients where histologic data is available, but its utilization may be considered in those with low-moderate level DSA and where an allograft biopsy is not accessible.

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“…For transplantations performed after September 2008, sera were retrospectively tested for the presence of anti-HLA antibodies with the LifeScreen Deluxe and Lifecodes single antigen beads class I and II (Immucor, GA, USA) in case of a positive screening. Based on a comparative study between both HLA antibody assays have a similar ability to detect anti-HLA antibodies at a mean fluorescence intensity cutoff of 4,000 ( 22 ), we decided to apply this cutoff. Donor-specific anti-HLA antibodies (DSA) were assigned for HLA-A/-B/-C/-DR/-DQ by comparing bead specificities with the donor HLA type on split level.…”

Section: Methodsmentioning

confidence: 99%

Association Between Promoter Polymorphisms in CD46 and CD59 in Kidney Donors and Transplant Outcome

et al. 2018

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Complement regulating proteins, including CD46, CD55, and CD59, protect cells against self-damage. Because of their expression on the donor endothelium, they are hypothesized to be involved in accommodation. Polymorphisms in their promoter regions may affect their expression. The aim of this study was to investigate if donor polymorphisms in complement regulating proteins influence kidney transplant outcomes. We included 306 kidney transplantations between 2005 and 2010. Five polymorphisms in the promoters of CD46, CD55, and CD59 were genotyped. A CD59 promoter polymorphism (rs147788946) in donors was associated with a lower 1-year rejection-free survival [adjusted hazard ratio (aHR) 2.18, 95% CI 1.12–4.24] and a trend toward impaired 5-year graft survival (p = 0.08). Patients receiving a kidney with at least one G allele for the CD46 promoter polymorphism rs2796267 (A/G) showed a lower rejection-free survival, though this became borderline significant after adjustment for potential confounders (aHR 1.87, 95% CI 0.96–3.65). A second CD46 promoter polymorphism (rs2796268, A/G), was also associated with a lower freedom from acute rejection in the presence of at least one G allele (aHR 1.95, 95% CI 1.03–3.68). Finally, the combined presence of both favorable genotypes of rs2796267 and rs147788946 had an additional protective effect both on acute rejection (p = 0.006) and graft survival (p = 0.03). These findings could help to identify patients who could benefit from intensified immunosuppressive therapy or novel complement inhibitory therapeutics.

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Comparative Assessment of Anti-HLA Antibodies Using Two Commercially Available Luminex-Based Assays

Abstract: Supplemental digital content is available in the text.

Cited by 20 publications

References 17 publications

Low incidence of IgA isotype of HLA antibodies in alloantigen exposed individuals

Low incidence of IgA isotype of HLA antibodies in alloantigen exposed individuals

Clinical utility of complement-dependent C3d assay in kidney recipients presenting with late allograft dysfunction

Association Between Promoter Polymorphisms in CD46 and CD59 in Kidney Donors and Transplant Outcome

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