Anti-HLA-B7/HLA-B44 strong cross immunoreactivity observed in flow cytometry HLA-B27 immunotyping

Area of Science:

• Immunogenetics
• Clinical Diagnostics
• Flow Cytometry

Background:

• Human leukocyte antigen (HLA) typing is crucial for clinical diagnostics.
• Cross-reactivities within the HLA-B7 related Cross-Reactive Group (B7CREG) can complicate HLA-B27 and HLA-B7 differentiation.
• Existing CE-IVD flow cytometry kits using double monoclonal antibodies (mAb) may yield complex results.

Purpose of the Study:

• To evaluate the performance of a specific HLA-B27 flow cytometry kit in distinguishing HLA-B27 and HLA-B7.
• To investigate unexpected cross-reactivities observed during routine HLA typing.
• To assess the reliability of HLA-B27 identification using current flow cytometry methods.

Main Methods:

• Analysis of 466 consecutive cases using the HLA-B27 IOTest™ kit on a Navios™ cytometer.
• Comparison of flow cytometry results with patient genotypes.
• Phenotypic analysis of anti-HLA-B7 and anti-HLA-B27 antibody labeling.

Main Results:

• Clear identification of expected HLA-B27-/HLA-B7- and HLA-B27+/HLA-B7- haplotypes.
• Observed three distinct phenotypes for anti-HLA-B27 labeling in patients positive for anti-HLA-B7, including cross-reactivity within B7CREG and unexpected reactions.
• Identified potential cross-reactivity of anti-HLA-B7 to HLA-B44 outside B7CREG and anti-HLA-B27 to other molecules.
• A 'grey zone' of intermediate intensity suggests challenges in definitively excluding HLA-B27 in some samples.

Conclusions:

• The study highlights significant and unexpected cross-reactivities in HLA-B27 and HLA-B7 typing by flow cytometry.
• Current mAb-based kits may not fully resolve complex HLA antigen expressions due to cross-reactivity beyond B7CREG.
• Further comparative studies are needed to refine HLA-B27 detection methods and establish reliable diagnostic criteria.