Analysis of human alpha-thrombin by hydrophobic interaction high-performance liquid chromatography.
The major active form of human thrombin, alpha-thrombin, was analyzed by hydrophobic interaction high-performance liquid chromatography (HIC-HPLC). The chromatographic system included a polymeric phenyl column and elution was performed by a gradient, 2-0M sodium chloride (5-20 min). Total analysis time was 30 min per injection. By this method, a good resolution between alpha-thrombin and the proteolytically modified thrombin forms, beta- and gamma-thrombin, was obtained. In addition, the thrombin preforms, prothrombin, prethrombin 1, and prethrombin 2, were also resolved from alpha-thrombin in the system. The results from the HIC method were compared to those obtained from non-reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis. By this high-resolution chromatographic method, the rapid analysis of purified alpha-thrombin is possible.
Karlsson
Göran
G
Plasma R&D, Octapharma AB, SE-112 75 Stockholm, Sweden. goran.karlsson@biovitrum.com
D002846:Chromatography, Affinity / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D013917:Thrombin / Q000032:analysis* / Q000737:chemistry* / Q000302:isolation & purification
D013917:Thrombin