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Reassessment of T Lymphocytes Crossmatches Results Prediction With Luminex Class I Single Antigen Flow Beads Assay

Visentin, Jonathan PharmD; Bachelet, Thomas MD; Borg, Cécile PharmD; Franchini, Nicolas PharmD; Nong, Thoa; Lee, Jar-How PhD; Couzi, Lionel MD; Merville, Pierre MD; Guidicelli, Gwendaline PharmD; Taupin, Jean-Luc PharmD

doi: 10.1097/TP.0000000000001239
Original Clinical Science—General

Background In virtual crossmatch (XM) strategies, a correct anticipation of XM results is required for appropriately allocating organs. We reassessed the ability to predict T lymphocyte flow cytometry and complement dependent cytotoxicity XM results with the mean fluorescence intensity (MFI) in Luminex class I single antigen flow beads (SAFB) assay, after correction of complement interference and exclusion of antidenatured HLA antibodies.

Methods Among 432 XM with T lymphocytes (T-XM), 407 were analyzed after exclusion of antidenatured HLA antibodies. Only ethylenediaminetetraacetic acid-treated serum MFI was considered. Only 1 cellular target HLA antigen for the serum was expressed in 238 cases, 209 and 29 being heterozygous and homozygous for this antigen, respectively. For the remaining 169 cases, at least 2 antigens were recognized. Single antigen flow bead MFI thresholds allowing XM positivity to be predicted were calculated with receiver operating characteristic curves.

Results T-XM results were tightly associated with SAFB MFI values. Anti-HLA-A and anti-HLA-B antibodies behaved similarly. Prediction and sensitivity SAFB MFI thresholds were determined, respectively, assessing the highest sensitivity/specificity ratio and at least 95% sensitivity for predicting T-XM positivity. Both were slightly lower for HLA-B than for HLA-A, whereas anti-HLA-Cw antibodies induced random XM results. Both thresholds were only slightly diminished for homozygous and for multiple HLA targets, considering the immunodominant, but not the sum, of antibodies MFI.

Conclusions Antibodies against HLA-A, -B, or -Cw behave differently. A homozygous HLA target does not trigger a twice higher XM positivity. Multiple antibodies are better evaluated through the immunodominant DSA MFI than through the sum of DSA MFI.

In the present study, the authors assess the correlations between Luminex class I single antigen flow beads assay (MFI) and T cell crossmatches in kidney transplant recipients after correction for complement interference and anti-denatured HLA antibodies. Prediction is good for anti-HLA A, B but not C antibodies. The information given by the immunodominant DSA MFI is compared to the sum of various DSA MFIs. Supplemental digital content is available in the text.

1 Laboratoire d’Immunologie et Immunogénétique, Hôpital Pellegrin, CHU de Bordeaux, Bordeaux, France.

2 UMR 5164 CNRS, Université Bordeaux Segalen, Bordeaux, France.

3 One Lambda, Inc., Canoga Park, CA.

4 Service de Néphrologie, Transplantation, Dialyse, Hôpital Pellegrin, CHU de Bordeaux, Bordeaux, France.

Received 28 December 2015. Revision received 14 March 2016.

Accepted 14 March 2016.

T.N. and J.L. are employees of One Lambda Inc. No specific funding was obtained for this study. Some SAFB assays were performed in One Lambda’s research department as a scientific collaboration, with reagents and equipments provided by One Lambda Inc and by staff people employed by One Lambda Inc. The other authors of this article declare no conflicts of interest.

J.T., J.V., and G.G. contributed to the design of the study. J.T. and J.V. participated in the writing of the article. J.V., T.B., T.N., J.L., N.F., and C.B. participated in the performance of the research. J.V., T.B., and J.T. participated in data analysis. J.M., P.M., and L.C. were involved in critical revision of the article.

Correspondence: Jean-Luc Taupin, PharmD, PhD, Laboratoire d'Immunologie et Histocompatibilité, Hôpital Saint-Louis APHP 1, avenue Claude Vellefaux 75475 Paris Cedex 10, France. (

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