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研究细胞动力学,对于基础和临床医学均有重要意义,国外已将胃肠道细胞动力学作为胃肠疾病的基础理论研究课题之一。在胃肠内分泌细胞动力学研究方面,近年来已有报道,应用体内注射放射自显影术和免疫组织化学相结合的方法来研究胃泌素细胞,但应用离体培养放射自显影术,对G细胞的观察,还甚少见。本实验是将离体培养放射自显影术与免疫组织化学法相结合,来显示标记G细胞。方法步骤如下:1.体外单次脉冲标记:(1)在基本无菌操作下,取胃幽门窦粘膜组织块,长2-3mm、宽约1mm,放入5ml链霉素灭菌瓶内,瓶中含有2mlEagle液(PH7.2-7.4),液中加有~3H-TdR 4μCi和10%小牛血清;培养液中还加入两只搅拌棒。橡皮塞封盖,用铁丝环夹加固。通过橡皮盖向瓶内注入空气3ml,以便促进~3H—TdR参入DNA中。(2)于37℃—37.5℃,经磁力搅拌器低速搅拌;孵育1小时,培养完毕,抽出空气;启盖,吸去培养液,换用Hank盐
Research on cell kinetics is of great importance to basic and clinical medicine. Gastrointestinal cell kinetics has been one of the basic theoretical research topics of gastrointestinal diseases in foreign countries. In the study of gastrointestinal endocrine cell kinetics, it has been reported in recent years that gastrin cells should be studied by in vivo injection of autoradiography combined with immunohistochemistry. However, in vitro culture of G Cell observation is still rare. This experiment is a combination of in vitro cultured autoradiography and immunohistochemistry to display labeled G cells. Method steps are as follows: 1. In vitro single-pulse labeling: (1) under basic aseptic operation, take gastric gland sinus mucosa tissue block, length 2-3mm, width 1mm, put 5ml streptomycin sterilization bottle, The bottle contains 2ml Eagle fluid (PH7.2-7.4), the solution was added ~ 3H-TdR 4μCi and 10% fetal bovine serum; also added to the culture medium two stirring rods. Rubber stopper cover, wire ring clamp reinforcement. The bottle was filled with 3 ml of air through a rubber cap to facilitate the incorporation of ~ 3H-TdR into the DNA. (2) at 37 ℃ -37.5 ℃, low speed stirring by a magnetic stirrer; incubated for 1 hour, the culture is completed, the air was extracted;