Objective:To establish a method for the isolation and purification of mice pancreatic islets during neonatal period. Methods:Pancreases of mice at neonatal stage were injected and digested by collagenase Ⅴ. The yield of islets was compared among different digestion time and hand-picked under microscope or Histopaque-1077 purification. The islets were stained with anti-glucagon antibody and anti-insulin antibody,and then identified by flow cytometry. Results:The optimal digestion time of islets from 1-week and 3-week old mice was 23 min (38.11 ± 2.78/pancreas) and 25 min (66.06 ± 2.61/pancreas). The diameters of most islets were less than 50 -滋m at 1 w (P < 0.01),and between 50~99 -滋m at 3 w (P < 0.01). The yield of islets purified by hand-picked under microscope was much higher than that of Histopaque-1077 (P < 0.05). The purity and survival rate of islets were above 90%,and the ratio of β and α cells within mice islets changed during neonatal period (P < 0.05). Conclusion:Local injection and digestion of pancreas with collagenase Ⅴ and hand-picked under microscope is an effective method for mice islet isolation during neonatal period,and the digestion time and purification methods are important. [Key words] neonatal period;mice;pancreatic islets;isolation and purification