Computer-aided immunofluorescence microscopy (CAIFM) for ANCA diagnostics

Introduction .– Screening by indirect immunofluorescence tests (IIFT) is the gold standard to detect anti-neutrophil cytoplasmic antibodies (ANCA). EUROPattern is a CAIFM system of microscope and software for evaluation and data management to standardize the reporting for IF assay (IFA) tests. Beyond recommended diagnostics with ethanol-fixed neutrophils, we provide a test system with 5 additional substrates for more precise diagnostics of ANCA-associated vasculitis (AAV).

Methods .– In one reaction field, three BIOCHIPs with cell substrates and three with purified antigen spots are combined: ethanol-fixed neutrophils, formalin-fixed neutrophils, an ethanol-fixed mixture of neutrophils and HEp-2 cells, and microdots of proteinase 3 (PR3), myeloperoxidase (MPO) and glomerular basement membrane (GBM).

A magazine holds up to 50 or 100 slides with 500 or 1000 reaction fields, and a camera takes photos of the reactions. Triggering, focusing and image recording is performed automatically or interactively using EUROLabOffice.

Results .– EUROLabOffice connects with the Laboratory Information Management System (LIMS), further liquid handling devices and manages data and test results. IIF images can be adjusted via a graphical user interface. Results are pre-classified as positive and negative. The user can zoom images, revise classifications, assign patterns, edit titers and results and access the patient history.

Discussion .– EUROPattern supports experts by simplifying and accelerating the laboratory workflow: data are processed consistently and traceable, automatic image recording enables fast archiving, the graphical user interface simplifies operators’ work and allows evaluating results with clean hands in an office instead of a dark room.

The automatic pre-classification is in good correlation with manual reading.

The 6-BIOCHIP mosaic achieves a broad screening with high sensitivity and specificity, even in the presence of interfering antibodies.

Conclusion .– The presented CAIFM system establishes a standardized process for ANCA evaluation.