Biosensor for diagnosing Factor V Leiden, a single amino acid mutated abnormality of Factor V.
Factor V Leiden (FVL) is an abnormality with a single amino acid mutation of Factor V (FV) and is the most common, hereditary blood coagulation disorder. FVL is currently diagnosed by DNA analysis, which takes a long assay time, high cost, and a specially trained person. We are developing a rapid, accurate, and cost-effective biosensing system to quantify both FV and FVL in blood plasma, to diagnose FVL and also to evaluate the seriousness of the disease status. This system is based on a sandwich immuno-reaction on an optical fiber. To produce the monoclonal antibody against only FV or only FVL without cross-reacting with the other molecule and with a higher probability, a 20 amino acid sequence (20-mer) of FV or FVL around the mutation region was injected into mice and then hybridoma cell lines specific to each 20-mer were selected. When these antibodies were tested with native FV or FVL molecules, they were found to be cross-reacting with the other molecules, but some with higher affinity to FV (FV preferred) and some to FVL (FVL preferred). Using these antibodies, two different sensors were developed: FV preferred and FVL preferred sensors. These two sensors allowed us to quantify FV and FVL in plasma with a maximum error of 4%. The plasma levels of both molecules provide us not only FVL diagnosis but also the level of the seriousness. The same principles may be used for developing diagnostic tools for other diseases with a single point mutation.
Ren
Yongjie
Y
Department of Chemical Engineering, University of Louisville, Louisville, KY 40292, USA.
Rezania
Samin
S
Kang
Kyung A
KA
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D015374:Biosensing Techniques / Q000191:economics / Q000295:instrumentation / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay; D005165:Factor V / Q000032:analysis / Q000235:genetics*; D006720:Homozygote; D006801:Humans; D006825:Hybridomas / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D017354:Point Mutation; D013997:Time Factors
D000911:Antibodies, Monoclonal; C095381:factor V Leiden; D005165:Factor V