title: Identification of a dysfibrinogen, the substitution of gamma308Asn(AAT) to Lys(AAG), using coagulation tests, immunoblot analysis, and allele-specific polymerase chain reaction.
abstract: During the past 3 years, we encountered 12 new cases suspected of being dysfibrinogenemias, via plasma coagulation screening tests, which included determination of fibrinogen concentration both by thrombin time and immunologic methods. We performed sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions and immunoblot analysis for these plasma fibrinogens. We identified two cases that were characterized by two distinct gamma-chain bands, similar to previous observations with Matsumoto-II (the substitution of gamma308Asn to Lys). Therefore, in order to identify the gamma308Lys variant easily and rapidly, we established an allele-specific polymerase chain reaction (AS-PCR). AS-PCR results indicated that the two cases were indeed heterozygous for the gamma308Lys variant; ten other cases were negative for this mutation. In conclusion, the ratio of fibrinogen concentrations determined by functional and antigenic methods in combination with the immunoblot analysis made these cases attractive for identifying the gamma308Lys mutation. The AS-PCR method proved to be a useful procedure to identify the gamma308K mutation.
authors.affiliation: Laboratory of Clinical Chemistry, Department of Medical Technology, School of Allied Medical Sciences, Shinshu University, 3-1-1 Asahi, Matsumoto, Japan.
authors.forename: F
authors.initials: F
authors.lastname: Terasawa
authors.forename: K
authors.initials: K
authors.lastname: Fujita
authors.forename: M
authors.initials: M
authors.lastname: Tozuka
authors.forename: H
authors.initials: H
authors.lastname: Ota
authors.forename: T
authors.initials: T
authors.lastname: Katsuyama
authors.forename: N
authors.initials: N
authors.lastname: Okumura
mesh_terms: D000483:Alleles; D019943:Amino Acid Substitution; D001216:Asparagine / Q000737:chemistry*; D001483:Base Sequence; D001780:Blood Coagulation Tests; D015153:Blotting, Western; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D015241:Fibrinogens, Abnormal / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008239:Lysine / Q000737:chemistry*; D008297:Male; D016133:Polymerase Chain Reaction
chemical_list: D017931:DNA Primers; D015241:Fibrinogens, Abnormal; D001216:Asparagine; D008239:Lysine