Electrophoretic analysis of endotoxin-activated gelation reaction of Carcinoscorpius rotundicauda amoebocyte lysate.
Electrophoretic analysis of a 60 min reaction between E. coli endotoxin and the amoebocyte lysate showed that the coagulation reaction was complete by 15 min, with the conversion of coagulogen (21 kDa) to coagulin (17 kDa). Coincident with this observation was the maximal activities at 15 min, of Factor C and proclotting enzyme. On agitation of the coagulin gel clots, bioactive endotoxin was recovered. Densitometric scan of the electrophoretically-resolved proteins showed that the sum of coagulogen and coagulin remained almost constant at various time intervals of the coagulation reaction. Electrophoresis serves as a convincing and visually discernible method of studying the kinetics of coagulation, and defining the onset and completion of gelation. Furthermore, it is a useful means of examining the integrity of fresh lysate preparations based on the presence or absence of the 17 kDa coagulin band.
Ho
B
B
Department of Microbiology, National University of Singapore, Kent Ridge.
Kim
J C
JC
Ding
J L
JL
D000818:Animals; D060829:Arthropod Proteins; D001798:Blood Proteins / Q000737:chemistry* / Q000378:metabolism; D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins / Q000737:chemistry*; D004792:Enzyme Precursors / Q000378:metabolism; D004926:Escherichia coli; D006737:Horseshoe Crabs; D012697:Serine Endopeptidases / Q000378:metabolism; D013925:Thromboplastin / Q000737:chemistry* / Q000378:metabolism
D060829:Arthropod Proteins; D001798:Blood Proteins; D004731:Endotoxins; D004792:Enzyme Precursors; C013445:coagulogen; D013925:Thromboplastin; D012697:Serine Endopeptidases; C057151:Limulus clotting factor C