An analysis of the role of collagenase and protease in the enzymatic dissociation of the rat pancreas for islet isolation.

摘要: Crude Clostridium histolyticum collagenase is widely used for the enzymatic degradation of pancreatic extracellular matrix in order to isolate islets Langerhans. The variable composition crude collagenases a critical issue which contributes poor reproducibility islet isolation procedures. In this study, separate contributions and protease process were analysed by testing various combinations purified rat pancreas dissociations under conditions eliminated all other proteolytic activity. Under these conditions, complete tissue dissociation required 99±10 min, whereas increasing amounts progressively reduced time minimum 36±1 min. Histochemical analysis showed that enhanced four major components matrix: collagen was degraded more completely, while proteoglycans, glycoproteins elastin at higher rate. Pancreas present, strictly controlled resulted high yield viable islets: 4.2–5.0 μl volume (3,300–3,800 islets) isolated per g presence or low concentration, respectively. Prolonged dramatic decrease correlated with observation enzyme accelerated disintegration. It concluded collagenase-induced facilitated protease. Our study shows yields can be obtained provided exposure limited.