Baranwal Ajay Kumar, Bhat Deepali Krishan, Goswami Sanjeev, Agarwal Sanjay Kumar, Kaur Gurvinder, Kaur Jasmeet, Mehra Narinder
Department of Transplant Immunology & Immunogenetics, All India Institute of Medical Sciences, New Delhi, India.
Department of Nephrology, All India Institute of Medical Sciences, New Delhi,, India.
Indian J Med Res. 2017 Feb;145(2):222-228. doi: 10.4103/ijmr.IJMR_222_16.
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.
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.
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.
供体人类白细胞抗原（HLA）特异性抗体在移植排斥和移植失败中起关键作用。近年来，其检测技术有了显著发展，从传统的基于细胞的检测方法到基于Luminex平台的先进固相系统，灵敏度和特异性不断提高。对于这些方法单独使用或联合使用的常规应用，目前仍未达成共识。本研究的目的是探索在Luminex平台上检测到的接近准确的平均荧光强度（MFI）截断值，以预测基于细胞的交叉配型结果的强度。
采用补体依赖细胞毒性试验（CDC）和流式交叉配型（FCXM）方法，检测116例等待移植的原发性肾移植受者血清样本中抗供体抗体的存在情况，并以相应供体作为对照，同时使用敏感的单抗原微珠（SAB）检测法检测HLA供体特异性抗体（DSA）。
MFI值<1000的HLA I类DSA患者，T细胞FCXM或CDC交叉配型均为阴性；MFI值在1000至3000之间的患者中，54%的FCXM为阳性，但CDC法检测均为阴性。然而，MFI值>3000的患者中，95%的FCXM为阳性。此外，MFI值在3000至5000之间的患者中，只有36%的CDC交叉配型为阳性，而MFI>7000的患者中90%为阳性。
基于Luminex SAB检测法的MFI截断值为3000时，与FCXM阳性显著相关；MFI值≥7000时，预测CDC交叉配型为阳性。作为CDC和FCXM检测的替代标志物获得的MFI截断值，将有助于解决不同基于细胞技术的局限性。
