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Investigating von Willebrand Factor Pathophysiology Using a Flow Chamber Model of von Willebrand Factor-platelet String Formation
Published on: August 14, 2017
[Establishment and Clinical Application of Flow Cytometric Immunobead Array in Detecting Plasma von Willebrand Factor
Bin Yan1, Yang He2, Shi-Qi Lu3
1Department of Clinical Laboratorial Medicine, Nanyang City Cential Hospital, Nanyang 473000, Henan Province, China,The First Affiliated Hospital of Soochow University, Jiangsu Institute of Hematology, Key Laboratory of Thrombosis and Hemostasis of Ministry of Health, Suzhou 215006, Jiangsu Province, China.
Insights
A new flow cytometric immunobead array (FCIA) accurately detects von Willebrand factor antigen (vWF:Ag). Elevated vWF:Ag levels in ischemic stroke patients predict poor prognosis and recovery.
Area of Science:
- Biochemistry
- Immunology
- Clinical Diagnostics
Background:
- Von Willebrand factor (vWF) is crucial in hemostasis and thrombosis.
- Elevated vWF levels are associated with ischemic stroke (IS) risk and prognosis.
- Accurate and efficient detection of vWF:Ag is essential for clinical assessment.
Purpose of the Study:
- To develop and validate a novel flow cytometric immunobead array (FCIA) for plasma vWF:Ag detection.
- To assess the clinical utility of FCIA in predicting prognosis for IS patients.
Main Methods:
- Developed an FCIA assay using anti-vWF antibodies and flow cytometry.
- Validated FCIA against ELISA using von Willebrand disease (vWD) and control samples.
- Measured plasma vWF:Ag in IS patients and correlated with 2-year prognosis.
Main Results:
- FCIA demonstrated excellent linearity (R2=0.99) and good agreement with ELISA (slope=0.97, bias=1.12%).
- FCIA is faster and more sensitive than ELISA, particularly for low vWF:Ag levels.
- IS patients showed significantly higher vWF:Ag, vWF:GPIbR, and vWF:CB levels compared to healthy controls (P<0.01).
Conclusions:
- FCIA is a sensitive, rapid, and effective method for vWF:Ag detection, complementing ELISA and improving type 3 vWD diagnosis.
- Elevated plasma vWF:Ag in IS patients is a significant indicator of poor prognosis and impaired functional recovery.
Objective:
To establish a novel flow cytometric immunobead array (FCIA) for detecting plasma von Willebrand factor antigen (vWF:Ag), and to analyze the clinical value of FCIA in predicting the prognosis of patients with ischemic stroke (IS).
Methods:
Anti-human vWF monoclonal antibody SZ29 IgG was coated on microspheres overnight, the diluted plasma was added after blocking, then incubated with FITC-conjugated sheep-anti-human vWF IgG polyclonal antibody, and finally detected by flow cytometry. The plasma vWF in 21 case of von Willebrand disease (vWD) and 105 controls (CTL) were detected by FCIA and ELISA, so as to carry out methodological assessment. Plasma vWF:Ag of 61 IS patients was detected by FCIA and the data of prognosis followed-up for 2-year were collected.
Results:
The linear fitting of FCIA was good (R2=0.99) without significant difference between FCIA and ELISA. The Bland-Altman bias was 1.12% with 95% limits of agreement that spanned from -45.06% to 47.30%, and the slope of the linear regression was 0.97 (r=0.86, P＜0.01). Importantly, the FCIA method was faster than ELISA, and superior to the ELISA in the detection of low levels of vWF:Ag. The levels of vWF:Ag, vWF:GPIbR and vWF:CB in IS patients were significantly higher than those in healthy controls (Z=8.36, 8.71, 6.22, respectively, P＜0.01).
Conclusion:
The FCIA for detecting plasma vWF:Ag is not only an effective supplement to ELISA, but also the efficiency is faster and more sensitive, thus improves the diagnosis of type 3 vWD. Elevated levels of vWF: Ag in IS patients indicate the poor recovery of daily activities and prognosis.
