Comparative Study Of Separation And Purification Of Different Day-old Changbai Pig Pancreatic Islet

Islet cell transplantation is one of the most effective therapeutic for the cure of Type I diaberes. The study of islet allotransplantation for solving islet source shortage was research focus to cure Type I diaberes. Thus, pig islets were considered to the optimum source of Human islet donor. The study discussed technology of pig pancreas islet isolation and purification. For all porcine islet isolation in this study, experimental animal was born was 1 day,60 days,120 days,165-180 days, adopt pancreiatic duct injection method, histopaque purification, enzyme linked immunosorbent assay and immunohistochemical technique to conduct experience.The result shows that:islet obtained amount and different collagenase digestion time were 0min,15min,30min,45min respectively, and each group was significant difference (P< 0.05), islet obtained amount with different concentration of collagenase were 1.0 mg/ml,1.5 mg/ml,2.0 mg/ml respectively and each group are significant difference (P< 0.05). During the same digestion time (15 min) and the day-old of 1 and 60 day to take collagen enzyme concentration was 1.0 mg/ml,120 and 165-180 day of collagen enzyme concentration of 1.5 mg/ml in the condition of purification,1 day-old porcine islet yield isolation was 3045 ±986 IEQ/g,2421+ 438 IEQ/g after purification, purity was 78%, the survival rate was 92%,2 months days porcine islet yield isolation was 2567±873 IEQ/g,1938 ± 362 IEQ/g after purification, the purity was 74%, survival rate was 94%,4 months days porcine islet cell yield isolation was 1969 ± 547 IEQ/g,1562 ± 236 IEQ/g after purification, the purity was 79%, survival rate was 90%,6 months days porcine islet yield isolation was 1758±125 IEQ/g,1485 ± 119 IEQ/g after purification, the purity was 84%,the survival rate was 84%. High glucose group of adult porcine insulin release was 788.56±77.53 mIU/IEQ, low glucose group was 158.37±28.44 mIU/IEQ. High glucose group of 165-180 day-old porcine insulin release was 679.58±68.69 mIU/IEQ, low glucose group was 102.43±23.12 mIU/IEQ. Porcine islet cells observed that 1 day-old group was 35±10 μm in diameter, and 60day-old porcine islet group was 62±20 μm in diameter, and 120 day-old porcine islet cells was 94±30 μm in diameter, and 165-180 day-old porcine islet group was 158±60 μm in diameter by HE staining. Porcine islet observed that 1 day-old group was 33±9μm in diameter, and 60day-old porcine islet cell group was 64±20 μm in diameter, and 120 day-old porcine islet was 91±32 μm in diameter, and 165-180 day-old porcine islet group was 143±54μm in diameter by DTZ staining.In this point, there was a difference in isolation and purification during different stage after borning. The optimum effect of 1 day-old and 60 day-old porcine islet cells isolation and purification for warm ischemia time was less than 10 min, and cold ischemia time was less than 30 min respectively, collagenase Ⅴ concentration was 1.0 mg/ml, digestion time for 15 minutes. The optimum effect of 120 days and 180 days porcine islet cells isolation and purification for warm ischemia time was less than 10 min, cold ischemia time was less than 30 min respectively, collagenase Ⅴ concentration was 1.5 mg/ml, digestion time for 15 minutes. The differentiative capacity, islet yield and islet release of piglet was superior to adult porcine.