Utilization of FPLC-purified bacterial collagenase for the isolation of cells from bone.
作者： Thomas J. Hefley
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摘要: Crude bacterial collagenase is essential for the enzymatic isolation of cells from membranous bone neonatal mouse calvaria. We have employed newly developed methodology fast protein liquid chromatography (FPLC) to separate and quantify isozymes so that their role in might be determined. FPLC resolved as many six peaks less than 30 min using a single anion exchange column separated into two classes. The Class I had preference substrate Azocoll, denatured collagen substrate, II Hexapeptide, synthetic substrate. Two preparations chromatographically purified (CGN-A CGN-B) were tested ability release viable bone. Both completely digested calvaria 120 min. total cell yield obtained with CGN-A was 0.34 million per calvarium. CGN-B 1.01 Each preparation analyzed FPLC. contained only isozymes, whereas mixture both isozymes. by then combined 4:1 ratio II: Utilization FPLC-separated increased 1.50 cells/calvarium. concluded there are combinations will digest extracellular matrix However, combination which favors result low rate destruction high yields.
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