Mean fluorescence intensity (MFI) values obtained from Luminex-based HLA antibody assays are used to determine the relative strength of HLA antibodies. However, MFI values can be inaccurate, especially when the patient's serum contains heterophile antibodies that are present in up to 40% of normal blood donors. Heterophile antibodies have been shown to cause false-positive results in Luminex-based immunoassays that use animal proteins such as bovine serum albumin and casein to block the reactive sites of the beads. However, these proteins provide a potential source of assay interference (false-positive results) as a result of heterophile antibody binding.

The aim of this study was to determine whether fetal bovine serum (FBS) would be an effective and reliable way to reduce heterophile antibody interference in Luminex-based HLA antibody assays. Toward this, we tested selected sera (n=5) containing heterophile antibodies by means of the LABScreen Single Antigen assays (One Lambda, Inc.). The sera were pre-treated with FBS (5% and 10%) and normal human serum (NHS) (5% and 10%), and their MFI values were compared to the ones from untreated samples. In addition, samples (n=20) without heterophile antibodies and containing well-defined HLA antibody profiles were also pre-treated with FBS (5% and 10%), and their MFI values were compared to the ones from untreated samples.

We show herein that pre-treatment with FBS (5% and 10%) eliminates heterophile antibody interference (MFI>1000) in several HLA class I (A29, A68, B67, and B76) and HLA class II (DR4, DR16, DRB3*02:02, DQA1*02:01/DQB1*03:01, and DPB1*19:01) specificities. On the contrary, NHS did not show any effect on the heterophile antibody interference in any of the samples. Moreover, FBS did not have any effect on the HLA antibody profile of sera displaying well-defined HLA antibody specificities.

The results of this study indicate that FBS-treatment is a cost-effective procedure to reduce heterophile antibody interference in Luminex-based HLA antibody assays. This interference can lead to a significant disadvantage for selected patients displaying heterophile antibody interference. Serum pre-treatment with FBS would prevent these patients from being inappropriately excluded from receiving an organ due to false-positive virtual crossmatch results.

CITATION INFORMATION: Jaramillo A., House J., Nelson L., Stoll S., Ridgley R., Khamash H. Serum Treatment with Fetal Bovine Serum Significantly Increases Luminex-Based HLA Antibody Assay Specificity by Reducing Heterophile Antibody Interference Am J Transplant. 2017;17 (suppl 3).

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