Fully automated immunoassay for quantitative determination of FXIII^[*]

Summary

Coagulation factor XIII (FXIII) is essential for clot stabilization. Deficiency of FXIII is associated with a risk of bleeding and impaired wound healing. Substitution therapy with FXIII remedies for patients with low plasma levels of FXIII requires diagnostic quantification of the factor before and during therapy. Here, we describe a prototype of a preliminary research immunoassay for quantification of FXIII antigen on automated coagulation instruments. The prototype assay is based on a monoclonal antibody (mAb) directed against FXIII A chain, whereas the mAbs are coupled to latex particles. FXIII in a plasma specimen causes agglutination of the latex particles, which can be quantified turbidimetrically. Performance data of the assay prototype processed on BCS^® XP and Sysmex^® CA-1500 instruments demonstrate a good correlation to the Berichrom^® factor XIII activity assay[1] from Siemens Healthcare Diagnostics (r = 0.94). Results: Comparability of instruments was excellent (r = 0.98). Coefficients of variation of total imprecision measurements ranged from 2.2 to 3.4%. Linearity was excellent over the range tested (12–121% FXIII). Analytical sensitivity was 0.51% FXIII on BCS XP and 0.44% FXIII on Sysmex CA-1500, respectively. No interference (> 10% bias) was observed with haemoglobin (up to 400 mg/dl), cholesterol (up to 300 mg/dl), bilirubin (up to 60 mg/dl) or triglycerides (up to 3000 mg/dl). Conclusion: The preliminary research assay prototype has the potential for excellent analytical sensitivity, precision, and dynamic range suitable to measure reliably FXIII antigen levels in human plasma.

Zusammenfassung

Gerinnungsfaktor XIII (FXIII) ist notwendig für die Stabilisierung eines Gerinnsels. Ein Mangel an FXIII geht einher mit erhöhtem Blutungsrisiko und gestörter Wundheilung. Die Substitutionstherapie mit FXIII-Präparaten bei Patienten mit niedrigen FXIII-Spiegeln erfordert eine diagnostische Quantifizierung von FXIII vor und während der Therapie. In dieser Arbeit wird der Prototyp eines vorläufigen Forschungstestes zur quantitativen Bestimmung von FXIII-Antigen auf automatisierten Gerinnungsanalyzern beschrieben. Der Prototyp basiert auf einem monoklonalen Antikörper (mAb) gegen die A-Kette des FXIII. Der mAb ist auf Latexpartikel gekoppelt. FXIII aus einer Plasmaprobe führt zur Agglutination dieser Latexpartikel, die turbidimetrisch gemessen werden kann. Ergebnisse: Vorläufige Leistungsdaten des Prototypen wurden auf den BCS^®-XPund Sysmex^®CA-1500-Systemen erhoben und zeigten eine gute Korrelation zu dem Berichrom^®-FaktorXIII-Aktivitätstest[1] von Siemens Healthcare Diagnostics (r = 0,94). Die Vergleichbarkeit der Ergebnisse zwischen den Systemen war hoch (r = 0,98). Die Variationskoeffizienten für die Bestimmung der Reproduzierbarkeit lagen zwischen 2,2 und 3,4%. Linearität war über den getesteten Bereich (12–121% FXIII) gegeben. Die analytische Sensitivität betrug 0,51% FXIII am BCS XP sowie 0,44% FXIII am Sysmex CA-1500. Keine Interferenz (> 10% Abweichung) wurde mit Hämoglobin (bis 400 mg/ dl), Cholesterol (bis 300 mg/dl), Bilirubin (bis 60 mg/dl) oder Triglyzeriden (bis 3000 mg/dl) beobachtet. Schlussfolgerung: Der Forschungsprototyptest weist das Potenzial für eine exzellente analytische Sensitivität, Präzision sowie einen dynamischen Messbereich auf, um FXIII-Antigen zuverlässig in menschlichen Plasmaproben zu quantifizieren.

Keywords

Bleeding - coagulation - factor XIII - factor XIII deficiency - immunoassay

Schlüsselwörter

Blutungen - Gerinnung - Faktor XIII - Faktor-XIIIMangel - Immunoassay

^* product under feasibility evaluation; not available for sale.

¹ not available for sale in the USA.

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