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Single Antigen Bead Testing: A novel testing methodology for detection of anti-HLA antibodies

The detection of anti-HLA antibodies helps to predict both the function and survival of transplant allografts.9 Most allograft damage is mediated though the activation of complement; C4d deposition is widely accepted as a marker for antibody-mediated rejection in renal allografts.1

Laboratory testing before transplantation includes cross matching donor cells (lymphocytes as surrogates of renal cells) with recipient serum. If a recipient has antibodies against a donor’s HLA type, the antibodies will bind to the donor’s cells and the crossmatch should be positive, whereas if the recipient does not have anti-HLA donor-specific antibodies, the crossmatch should be negative.

The transplant based on both a negative complement-dependent cytotoxicity (CDC) and the more sensitive flow cytometric crossmatch. However, both can be subject to interference, 2,3 and even
if both crossmatches are negative, early graft rejections still occur. Higher organ rejection rates occur among patients with donor specific anti-HLA antibodies prior to transplantation.4

The presence of preformed human leucocyte antigen (HLA) donor specific antibodies (DSA) is a major risk factor for early renal allograft rejection and graft loss [5,6]. Different techniques are currently available to detect anti-DSA in sera from patients waiting for a graft.

The standard complement-dependent cytotoxicity crossmatch (CDCXM) and some modification of this technique such as the anti-human globulin-augmented assay (AHG-CDCXM) have been the most widely used to detect anti-DSA before transplantation [7-9]. The widespread utilization of CDCXM against donor T lymphocytes has resulted in the almost complete elimination of hyperacute rejection [10-12].

The flow cytometric crossmatch (FCXM) is more sensitive than CDCXM [10–12]; nevertheless, this technique is not specific because it can detect antibodies directed against cell surface antigens irrelevant in the outcome of the graft. The issue of FCXM being a more sensitive but less specific technique when compared with CDCXM, leads to false-positive results, preventing the opportunity of a transplant in patients without relevant DSA.

The single-antigen bead flow Cytometry (SAFC) allows the determination of the presence of DSA comparing the HLA typing of the donor with the repertory of anti-HLA antibodies against recombinant HLA molecules in the serum of each patient (virtual XM)


The Luminex anti-HLA antibody detection assay is reportedly more sensitive and specific than either the CDC or flow cytometric crossmatches.4,13,14 In fact, some consider the Luminex antibody detection technique to be the new gold standard for identifying anti-HLA antibodies.5 This technique is a solid-phase assay in which purified HLA molecules (either of a single HLA type or a combination of types) are attached to beads. These molecules will bind to anti-HLA antibodies in the patient’s serum.4,14 Using single antigen technology, the Luminex technology can predict a patient’s sensitization to particular HLA types prior to transplantation without performing a physical CDC or flow cytometric crossmatch (termed a ‘virtual crossmatch’).

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