腫瘤藥物敏感性試驗（2）－－藥物敏感性試驗的方法學

Tumor Chemosensitivity Assay, Ⅱ: Methodology of ChemoSensitivity Assay

本文旨在介紹我們所用的體外腫瘤藥物敏感性試驗（microculture tetrazolium試驗，MTT試驗）的方法，並評價其可靠性。MTT試驗為一敏感、簡便、定量的比色測定法，已廣泛用作腫瘤藥物敏感性試驗。原理為活細胞的mitochondrial dehydrogenase enzyme可將黃色的試劑tetrazolium bromide轉化成深藍色的formazan，產生的formazan量與活細胞數成正比，然後用多皿掃描比色計讀數，即可瞭解生存的細胞數。本文通過以下二項試驗研究本法的可靠性：（1）對本組患者（53例）作了5種藥物（5Fu, mitomycin, adriamycin, cisplatin, methotrexate）的劑量－效應測定。發現細胞數與藥物濃度有高度相關（負相關），二者的相關係數統計上有顯著意義，顯示二者間有密切關連。（2）在13例患者對4種藥物（5Fu, mitomycin, adriamycin, cisplatin）作了二次試驗，將其結果進行比較。發現同一例患者對各種藥物所做二次測定的結果（ID50）非常接近，呈正相關，二者的相關係數有顯著意義。二次測定結果符合率（指二次均為敏感或均為耐藥）達96.2%。本文顯示MTT試驗為一可靠的腫瘤藥物敏感性試驗，具有較好的可重複性。

The purpose of the present study was to describe the methodology of our in vitro tumor chemosensitivity test (microculture tetrazolium test, MTT assay) and to evaluated the reliability of this assay.The MTT assay is a sensitive, quantitative and convenient colormetric assay and has been used widely as an in vitro tumor chemosensitivity test. The assay is based on the capacity of mitochondrial dehydrogenase enzymes in living cell, to convert the yellow tetrazolium bromide into a dark blue formazan product. The amount of formazan produced is directly proportional to the number of living cell. The technique permits the processing of a large number of samples using a multiwell scanning spectrometer.We investigated the reliability of the MTT assay by :(1) The dose-response relationship of five anticancer drugs (5FU, mitomycin, adriamycin, cisplatin and methotrexate) were determined in the present group (53 cases). The cell number showed a high correlation with the drug dose (negative correlation) and the correlation coefficients between the cell number and drug dose were statistically significant, indicating the signififcant correlation between these two parameters. 2. In 13 cases, the MTT assay for four anticancer drugs (5FU, mitomycin, adriamycin and cisplatin) was performed twice and the results were compared. The paired ID50 of various drugs obtained from these two tests showed a positive correlation and the correlation coefficients were highly significant. The rate of consistency between these two tests was 96.2% (both tests show sensitive or resistant results). The present study showed that the MTT assay is a reliable in vitro tumor chemosensitivity assay with excellent reproducibility.