PB0805 - Comparison of three ELISA assays for the detection and quantification of autoantibodies against complement factor H

Monday, June 26, 2023

18:30 – 19:30 ET

Room: Exhibition

Presenting Author(s)

Sandrine THOUZEAU, PhD (she/her/hers)

PhD student
Division of Hematology-Oncology, CHU Sainte-Justine, Université de Montréal
Montreal, Quebec, Canada

Co-Author(s)

Clémence Merlen, PhD (she/her/hers)

Research Associate
Division of Hematology-Oncology, CHU Sainte-Justine, Université de Montréal
Montreal, Quebec, Canada
EP

Emmanuelle Pépin

Research assistant
Division of Hematology-Oncology, CHU Sainte-Justine, Université de Montréal
Montreal, Quebec, Canada
LB

Laurine Boulanger

Ing student
Division of Hematology-Oncology, CHU Sainte-Justine, Université de Montréal
Montreal, Quebec, Canada
RY

Rayan Yousfi

Bachelor student
Division of Hematology-Oncology, CHU Sainte-Justine, Université de Montréal
Montreal, Quebec, Canada
GR

Georges-Etienne Rivard

Medical Director, Hemophilia Center, CHU Sainte-Justine
Division of Hematology-Oncology, Department of Pediatrics, Université de Montréal, CHU Sainte-Justine, Montréal, Quebec, Canada
Montreal, Quebec, Canada
ST

Stéphan Troyanov

Head of Department
Division of Nephrology, Hôpital du Sacré-Cœur, Université de Montréal
Montreal, Quebec, Canada
AB

Arnaud Bonnefoy

Associate Professor
Division of Hematology-Oncology, CHU Sainte-Justine, Université de Montréal
Montreal, Quebec, Canada

Background: The screening for autoantibodies against factor H (FH), a major regulator of the alternative complement cascade, is recommended for the diagnosis of autoimmune atypical hemolytic uremic syndrome (aHUS) and mandatory during the monitoring of associated immunosuppressive therapy.

Aims: Compare and evaluate three direct ELISA assays for the detection and quantification of anti-human FH immunoglobulin G.

Methods: For the purpose of the study, 78 citrated plasma samples from 51 patients referred to the hemostasis laboratory of the CHU Ste-Justine for suspicion of auto-immune aHUS were measured. The results of two commercial quantitative assays expressed in arbitrary units (AU/mL): Anti-Faktor H assay (GA Generic Assays, Dahlewitz, Germany) and CFH IgG ELISA kit (Abnova, Taipei, Taiwan) were compared to those obtained with a reference in-house direct ELISA titration assay, which included individual measurement of non-specific signal. Concordance between assays was evaluated using Pearson correlation for quantitative comparison, and Cohen’s kappa for categorical agreement.

Results: 34/78 samples were positive for anti-FH with the in-house assay. 43/78 were negative and 1/78 was in a grey zone. Five false positive samples were detected through non-specific signal correction. Positivity threshold were 3.7 AU/ml for the Generic Assays kit and 9.0 AU/ml for the Abnova kit. Global concordance was 71% between the three assays (24 positive and 31 negative samples in all three assays (figure 1). The in-house and Generic Assays methods had the best rate of agreement (85%), Cohen’s kappa (0.69) and Pearson’s correlation (r2 = 0.65) (Table 1).

Conclusion(s): Although good agreement can be obtained between anti-FH ELISAs, significant discrepancies remain that require caution in the interpretation of results. The introduction of an individual control to detect false positives or a confirmatory test with a different kit is strongly recommended.