离体肺组织切片的制备和孵育

Preparation and incubation of rat precision-cut lung slices

目的建立一种简便有效的离体肺组织切片孵育方法。方法利用肺组织精细切片技术制备肺组织切片 ,以 0 .5mLKrebs Henseleit(K H)缓冲液作为孵育液 ,在 37℃培养箱内分别孵育 1、2、3、4h ,测定肺片的四甲基偶氮唑盐 (MTT)比色、乳酸脱氢酶 (LDH)释放率和三磷酸腺苷 (ATP)含量 ,以反映肺组织活性。结果肺片在 37℃培养箱内孵育 1、2、3、4h后MTT比色、LDH释放率和ATP含量均无统计学差异 (P >0 .0 5 )。结论应用 0 .5mLKrebs Henseleit(K H)缓冲液作为孵育液、在 37℃培养箱对肺片进行孵育时组织活性没有明显变化 。

Objective To establish an effective but simple method for the incubation of precision-cut lung slices. Methods The rat precision-cut lung slices were prepared and incubated for 1, 2, 3, 4 h in 37℃ incubator taking Krebs-Henseleit(K-H)buffer as incubation liquid. The quantitative measurement of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenylt-etrazolium bromide (MTT) colorimetric assay and contents of ATP in incubated lung slices, the lactic dehydrogenase (LDH) releases in the incubated supernate of lung slices were detected as the indexes reflecting viability of lung slices. Results Absorbency of MTT colorimetric assay, ATP content and LDH release of lung slices incubated 1, 2, 3, 4 h in 37 ℃ incubator remained unchanged, respectively(P>0.05). Conclusion Precision-cut lung slices could be incubated effectively and simply in 37 ℃ incubator with unchanged viability, taking 0.5 mL Krebs-Henseleit (K-H) buffer as incubating liquid.