摘要
为研究牛乳腺组织冷冻保存的可行性,将牛乳腺组织剪成直径小于0.5 mm的组织块,以含20%NBS和10%DM-SO的DMEM/F12作为冷冻保护液,按如下程序进行冷冻保存:4℃平衡1 h,-20℃平衡2 h,-40℃平衡2 h,-80℃保存。1周后,复苏牛乳腺组织,检测组织的存活率,分离培养乳腺细胞,检测细胞的存活率,观察细胞生长状态。结果显示,复苏后的牛乳腺组织可重新贴壁生长,贴壁存活率为62.5%,经消化离散,可分离培养出乳腺细胞,细胞存活率为58.5%,培养24 h后细胞存活率为46.3%。随着培养时间延长,细胞活力恢复,第1代细胞存活率为93.2%,生长状态良好。该乳腺组织的冷冻方法操作简单、方便、快捷,且很好地保持了乳腺细胞的活力,为稳定和增加乳腺细胞的来源提供了很好的途径,为进一步研究乳腺的结构和发育提供了素材。
To investigate the feasibility of cryopreservation of bovine mammary tissue,bovine mammary tissue was cut into pieces with the diameter under 0.5 mm and frozen using cryoprotectant was DMEM/F12 including 20% NBS and 10% DMSO.The mammary tissue pieces were cryopreserved according to the following procedures： balanced at 4 ℃ for 1 h,-20 ℃ for 2 h,-40 ℃ for 2 h,and preservated at-80 ℃.After a week,recovered the mammary tissue and quantified the survival rate of tissue pieces.Cell viability was assessed on cell suspensions immediately after digestion and a 24 h culture period.The cells growth morphology was observed.It was found that recovery of bovine mammary tissue could adhere and grow,and the survival rate was 62.5%.After digesting,mammary cells could be isolated and the cell morphology from frozen-thawed samples was similar with fresh samples.The cells living rate was 58.5% when just isolated and it decreased to 46.3% after culture for 24 h.Long-term culture of mammary cells was carried out and a good growth status was offered.The mammary cells living rates of passages 1 increased to 93.2%.The frozen method of bovine mammary tissue was simple,convenient and fast,which could be used to maintaine mammary tissue viability,would stabilize and expand the source of mammary cells,and provide a stable material to study the structure and developmental biology of mammary gland.
出处
《中国畜牧兽医》
CAS
北大核心
2011年第5期42-45,共4页
China Animal Husbandry & Veterinary Medicine
基金
优质高效转基因肉牛新品种培育(2009ZX08007-005B)
关键词
牛
乳腺组织
冷冻保存
bovine
mammary tissue
cryopreservation
