Purification of rabbit C-reactive protein by affinity precipitation with thermosensitive polymer.
A method of purifying rabbit C-reactive protein (CRP) based on affinity coprecipitation with a thermosensitive polymer is developed. This polymer, which was obtained by conjugation of p-aminophenylphosphorylcholine (APPC) and poly-N-isopropylacrylamide derivative with reactivity to primary amino groups, exhibited a thermally reversible phase separation profile with a critical value at 32 degrees C and high affinity for CRP in the presence of Ca2+. When rabbit acute-phase serum was incubated with APPC-conjugated polymer in the presence of Ca2+ at 4 degrees C, only CRP was bound to this affinity polymer and coprecipitated by raising the temperature of the incubation mixture above 32 degrees C. After addition of cold EDTA solution to the precipitate followed by centrifugation at 38 degrees C, CRP was recovered from the polymer in more than 80% yield with no contaminants. The binding of CRP to APPC-conjugated polymer was dependent on concentrations of both CRP and the polymer.
Mori
S
S
School of Health Sciences, Okayama University, Japan.
Nakata
Y
Y
Endo
H
H
D000178:Acrylamides; D000210:Acute-Phase Reaction / Q000097:blood; D000818:Animals; D002097:C-Reactive Protein / Q000302:isolation & purification*; D011232:Chemical Precipitation; D004797:Enzyme-Linked Immunosorbent Assay; D004852:Epoxy Compounds; D006358:Hot Temperature; D008689:Methacrylates; D010767:Phosphorylcholine / Q000031:analogs & derivatives*; D011108:Polymers; D011485:Protein Binding; D011817:Rabbits
D000178:Acrylamides; D004852:Epoxy Compounds; D008689:Methacrylates; D011108:Polymers; C088774:4-aminophenylphosphorylcholine; D010767:Phosphorylcholine; D002097:C-Reactive Protein; C067295:N-isopropylacrylamide; C007870:glycidyl methacrylate