CCK-8法和MTT法评价医用抗菌不锈钢细胞毒性的比较研究

[摘要] 目的 通过MTT法和CCK-8法对医用抗菌不锈钢的细胞毒性进行比较研究,为其临床应用提供生物学证据。方法 将各组材料制备成15 mm×10 mm×3 mm的长方体试件,按样品表面积:溶液体积3 cm~2/ml的比例制备浸提液,通过倒置相差显微镜观察共培养24、48、72 h的细胞形态,并通过MTT比色法和CCK-8法得出细胞在培养24、48、72 h的吸光度(A)值,进而计算其RGR评价抗菌不锈钢的细胞毒性。结果 ①培养48 h后,随着作用时间的延长,阳性对照组细胞表现出明显的中毒形态,其余各组细胞生长旺盛,医用不锈钢及抗菌不锈钢组开始出现少量细胞核固缩。②MTT实验中,阴性对照组、不锈钢试件组及阳性对照组的A值呈递减趋势,但抗菌不锈钢和普通医用不锈钢的A值无显著差异。③CCK-8实验结果显示,各组A值按阴性对照组、抗菌不锈钢试件组、普通医用不锈钢对照组、阳性对照组的顺序呈递减趋势,差异有显著性。④实验中各组试件的细胞毒性均为0级。结论 ①抗菌不锈钢毒性等级与医用不锈钢相同,符合口腔正畸材料的临床应用要求。②CCK-8法较MTT法具有更高的灵敏度和可重复性,值得推广应用。

Comparative study on the cytotoxicity of antibiotic stainless steels by the methods of MTT and CCK-8

Objective To evaluate the cytotoxicity of antibiotic stainless steels by the methods of MTT and CCK-8.Mehtods Metal test samples were made into rectangular solids in size of 15 mm x 10 mm x 3 mm.Alloy leaching liquor was prepared with DMEM culture media according to the ratio of surface area and volume of culture solution(3 cm /ml).Growth of MG63 cells was observed under inverted phase contrast microscope;absorbance of cells cultured for 24 to 72 hours was detected using MTT and CCK-8 test;cytotoxicity of antibiotic stainless steels was evaluated.Results①After 48 hours’ culture,with the culture time increased,cytotoxicity was detected in the positive control group;cells in other groups were normal and grew well.A few of cells in stainless steels group showed karyopyknosis.②Although the A value of antibiotic stainless steel group was higher than that of stainless steel group from MTT results,there was no significant difference between the two groups.③There were significant differences among the four groups from CCK-8 results.The A value gradually decreased from the negative control group to the positive control group.④The level of cytotoxicity in each of the test groups was grade 0.Conclusions①The antibiotic stainless steel is in line with the requirement of its clinical application by the cytotoxicity test.②The CCK-8 method is more sensitive and accurate than MTT and worth using.