PB0070 - Fluorescence cross-correlation spectroscopy based measurement of von Willebrand factor multimer size for classifying patients with acquired von Willebrand syndrome

Background: Acquired von Willebrand syndrome (AVWS) is a complication of cardiovascular disease or mechanically assisted circulation, caused by shear-stress dependent breakage of von Willebrand factor (VWF) multimers. Conventionally, loss of VWF multimers is detected by multimer analysis, which is lengthy, qualitative and subjective in nature.

Aims: We developed a simple, rapid and quantitative immunoassay based on fluorescence cross-correlation spectroscopy (FCCS), for measuring VWF size changes [1]. We aim to investigate the potential of our assay for diagnosing AVWS.

Methods: Plasma VWF is bound to fluorescently labeled antibodies in situ, and losses of multimers are detected by measuring changes in the diffusion speed of the immunocomplex in solution. Diffusion measurements are normalized with the value of a traceable calibrator to obtain VWF size in percentage units. Plasma VWF sizes were measured for suspected AVWS patients classified using a VWF Large Molecular Weight index (LMWi) obtained by multimer analysis [2]. Written informed consent was obtained from all volunteers. The study was approved by the Institutional Review Boards of Tohoku University and Sysmex Corporation and funded by Sysmex Corporation.

Results: Plasma VWF sizes measured using our assay could differentiate between AVWS and non-AVWS cases with a high AUROC (> 0.8). Higher performance was observed in differentiating severe AVWS patients with remarkably reduced multimers as detailed in table 1. Also, VWF size showed a moderately positive correlation with LMWi.

Conclusion(s): Using 1 μl of plasma, less than 10 ng of antibodies per test, and 30 minutes to results we indicate that plasma VWF sizes measured using our FCCS-based immunoassay may be used to rapidly quantify changes in the multimeric status of VWF and potentially be used as a standardizable alternative for multimer pattern analysis after further validation.

References:
1. Ganguly, et al, 2023. Anal. Chem., 95, 21, 8172–8179.ss
2. Tamura, et al, 2015. J. Atheroscler. Thromb., 22, 11, 1115-1123.