title: Rapid determination of blood coagulation factor XIII activity using protein arrays for serodiagnosis of human plasma.
abstract: We developed a novel on-chip assay using protein arrays for quantitative and rapid analysis of blood coagulation factor XIII (FXIII) activity in human plasma. FXIII is activated by concerted action of thrombin and Ca(2+) and plays essential roles in hemostasis, angiogenesis, and wound healing. We fabricated protein arrays by immobilizing fibrinogen onto the 3-aminopropyltrimethoxysilane layer of well-type arrays and determined FXIII activity by analyzing biotinylated fibrinogen with Cy3-conjugated streptavidin. We determined optimal concentrations of Ca(2+), thrombin, and 5-(biotinamido)pentylamine (BAPA) for the on-chip activity assay, and the detection limit was 0.01 Lowey U/mL (9.9 pM). Using the on-chip activity assay, hepatocellular carcinoma patients (n = 24), but not hepatitis (n = 24) or liver cirrhosis patients (n = 41), had significantly lower FXIII activities (p < 0.001) than normal individuals (n = 41), indicating that FXIII activity is a possible diagnostic marker for hepatocellular carcinoma. In addition, we have successfully used this activity assay to reveal individual variations (37-57%, n = 65) in the inhibition rate of FXIII activity by isoniazid, the first-line antituberculosis agent. Thus, our optimized on-chip FXIII activity assay provides a quantitative and high-throughput approach to investigating the role(s) of FXIII in human diseases. Moreover, it has a strong potential to be applied toward FXIII-related personalized medicines.
authors.affiliation: Department of Molecular and Cellular Biochemistry and Institute of Medical Science, Kangwon National University School of Medicine, Chuncheon, Kangwon-do 200-701, South Korea.
authors.forename: Mi-Hye
authors.initials: MH
authors.lastname: Kwon
authors.forename: Deok-Hoon
authors.initials: DH
authors.lastname: Kong
authors.forename: Se-Hui
authors.initials: SH
authors.lastname: Jung
authors.forename: In-Bum
authors.initials: IB
authors.lastname: Suh
authors.forename: Young-Myeong
authors.initials: YM
authors.lastname: Kim
authors.forename: Kwon-Soo
authors.initials: KS
authors.lastname: Ha
mesh_terms: D000588:Amines / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000037:antagonists & inhibitors / Q000378:metabolism; D001710:Biotin / Q000031:analogs & derivatives / Q000378:metabolism; D001800:Blood Specimen Collection; D006528:Carcinoma, Hepatocellular / Q000097:blood / Q000175:diagnosis; D005176:Factor XIII / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism; D006505:Hepatitis / Q000097:blood / Q000175:diagnosis; D006801:Humans; D055767:Immobilized Proteins / Q000378:metabolism; D007538:Isoniazid / Q000494:pharmacology; D057230:Limit of Detection; D008103:Liver Cirrhosis / Q000097:blood / Q000175:diagnosis; D008113:Liver Neoplasms / Q000097:blood / Q000175:diagnosis; D040081:Protein Array Analysis; D015203:Reproducibility of Results; D012698:Serologic Tests / Q000379:methods*; D013997:Time Factors
chemical_list: D000588:Amines; D014408:Biomarkers, Tumor; D055767:Immobilized Proteins; C076022:5-(biotinamido)pentylamine; D001710:Biotin; D005340:Fibrinogen; D005176:Factor XIII; D007538:Isoniazid