Qualitative detection of the Marburg I alloenzyme of factor VII-activating protease by an immunoassay and its comparison to PCR testing.
BACKGROUND
The Marburg I (MRI) single nucleotide polymorphism (SNP) of the factor VII-activating protease (FSAP) gene has been associated with thrombophilia, thromboembolism, atherosclerosis, and the incidence and progression of carotid stenosis. At present, MRI SNP testing is mainly performed using costly nucleic acid analysis. The ratio between FSAP activity and antigen concentrations in citrated plasma has been used to assess the FSAP genotype.


METHODS
This article describes the development of a prototype ELISA for the detection of the MRI FSAP alloenzyme, and its correlation to FSAP genotypes to assess whether a positive MRI FSAP ELISA result may be used as a surrogate marker for the presence of the MRI SNP.


RESULTS
ELISA results were correlated with FSAP genotypes from 523 blood donors measured using PCR. Diagnostic sensitivity and specificity of the assay for determination of the genotype were 100% (95% confidence interval [CI]: 93.36-100) and 99.79% (95% CI: 98.80-99.96), respectively. Maximum run-to-run, within-run, and total coefficients of variation were 7.8%, 7.9%, and 9.9%, respectively. No cross-reactivities with homologues of the MRI FSAP alloenzyme were observed. Test performance was not affected by typical interfering compounds.


CONCLUSIONS
The data demonstrate that an immunoassay applying antibodies specific to the MRI FSAP alloenzyme can provide sufficiently accurate detection of the MRI SNP. This will significantly simplify MRI FSAP testing, particularly in large cohorts.
Schwarz
Herbert
H
Siemens Healthcare Diagnostics Products GmbH, Marburg, Germany.
Stephan
Sina
S
Sidelmann
Johannes J
JJ
Fischer
Bodo
B
Althaus
Harald
H
Hahn
Martin
M
Kappel
Andreas
A
Ehm
Matthias
M
Vitzthum
Frank
F
D002318:Cardiovascular Diseases / Q000235:genetics; D004789:Enzyme Activation; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000592:standards; D015942:Factor VIIa; D007118:Immunoassay; D007527:Isoenzymes / Q000097:blood; D016133:Polymerase Chain Reaction / Q000592:standards*; D020641:Polymorphism, Single Nucleotide; D012680:Sensitivity and Specificity; D012697:Serine Endopeptidases / Q000097:blood*
D007527:Isoenzymes; C433550:HABP2 protein, human; D012697:Serine Endopeptidases; D015942:Factor VIIa