激光扫描共聚焦显微镜观察细胞在丝素纤维织物上的生长情况

Observation grawth of cells on natural and regenerated silks fibronin with laser scanning confocal microscope

利用激光扫描共聚焦显微镜(laser scanning confocal microscope,LSCM)技术,用活细胞荧光示踪剂标记细胞,观察细胞在天然丝素纤维和再生丝素纤维上的生长情况。用含10μmol/L Cell-Tracker CM-DiI(Molecular Probes Company)的无血清RPMIl640培养液,室温标记V79、3T3和原代心肌细胞30min,接种于Glass Bottom Dish中,将两种受试材料分别放入Glass Bottom Dish中,继续培养。在加入受试材料后1、2、3、4、5,7天,利用LSCM技术,采用双通道同时扫描的方式,一个通道的激发波长为534nm,发射波长为540-650nm,另一个通道使用微分干涉差成像技术,动态观察细胞在再生丝素纤维受试材料的生长情况。实验发现V79细胞和3T3细胞在天然丝素纤维上是接种后5天明显增多,在再生丝素纤维上生长数量是接种后4天明显增多,而且比在天然丝素纤维生长的数量多。原代心肌细胞在天然和再生丝素纤维上生长得非常少。

Objective：Growth of cells on natural and regenerated silk fibroin was investigated with LSCM. Methods： V79, 3T3 and myocardium cells were loaded with 10μmol/L Cell- Tracker CM- Dil （Molecular Probes Company） and cultured in the Glass Bottom Dish. On the ld, 2d, 3d , 4d, 5d, 7d after adding natural and regenerated silk fibroin, the growth of cells on the silk was observed with detector 1 （534nm of optimum excitation wavelength） and detector 2 （differential interference contrast measurement, DIC） by laser scan confocal microscope. Result：The numbers of V79 and 3T3 cells on the regenerated silk on 4d were increased, and cells on the natural silk were increased on the 5d. The numbers of cells on the regenerated silk were more than those on the natural silk. However, myocardium cells on the regenerated and natural silk were less.