ELISA-Based Crossmatching Allowing the Detection of Emerging Donor-Specific Anti-HLA Antibodies through the Use of Stored Donors’ Cell Lysates

Figure 1

Flow diagram of the AMS-ELISA shown for the detection of HLA class I molecules. (a) Binding of the donor’s solubilized HLA class I molecules by monoclonal capture antibodies recognizing a monomorphic epitope on HLA class I molecules. (b) Binding of the donor-specific anti-HLA antibodies out of the recipient’s serum to the HLA molecules of the donor. (c) Binding of alkaline phosphatase conjugated secondary antibodies to the recipient’s bound donor-specific anti-HLA class I antibodies and subsequent color reaction. The original protocol was modified by substituting the human IgG-specific antibody by a human IgG/M/A-specific secondary antibody. (d) Lysate control using an alkaline phosphatase conjugated monoclonal detection antibody directed against a second monomorphic epitope to confirm the immobilization of a sufficient amount of HLA molecules by the solid-phase-bound capture antibody. The AMS-ELISA variant for the identification of donor-specific antibodies directed against HLA class II molecules is correspondingly designed.

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