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Title: Comparison of Three Non-Radioactive Assays for Measurement of Cell-Mediated Immunity
Other Titles: 比較三種偵測細胞免疫反應之非放射線法
Authors: 林大盛
Issue Date: Dec-2006
Publisher: 國立臺灣師範大學生命科學學系
Department of Life Science, NTNU
Abstract: 細胞反應可以放射線或非放射線法來衡量。雖然目前有非放射線法可用,但由於放射線法之靈敏度及可信度相當高,故仍常被使用。雖然如此,放射線危險性及其廢棄物是否安全處置常引起大眾關心。本研究比較三種非放射線法偵測ICR小鼠淋巴結細胞對萃取自CFW鼠sarcoma-180(S-180)全細胞、膜、及其內部成份之可溶性抗原反應之靈敏度。在細胞-小珠粒共同培養系統,根據細胞聚集之程度與形式,記錄為細胞反應指數。氨基黑素試驗與3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide(MTT)試驗乃利用其呈色反應定量細胞受刺激反應增殖程度。結果一致顯示,取自S-180免疫小鼠之淋巴結細胞顯著的對S-180抗原有反應,但對不相干的刺激物-牛血清白蛋白卻沒反應。在S-180三種抗原中,全細胞抗原對於S-180致敏細胞很明顯是最好的刺激原。在三種測試法,MTT試驗的靈敏度最好,因其可顯著區別對三種S-180抗原的反應。細胞刺激強度最高的是S-180全細胞抗原,其次是內部抗原,最低的是膜抗原。氨基黑素試驗僅稍顯著的區分對S-180內部抗原及膜抗原的反應。在細胞-小珠粒共同培養系統,當免疫細胞和S-180全細胞抗原附著之小珠粒一起培養時,細胞反應指數達到最高。雖然如此,當使用內部或膜抗原附著之小珠粒時,並沒顯著差別。總而言之,在測量細胞受刺激反應增殖程度,非放射線法安全且相當靈敏。這些測試法可彼此互補使用,以測量細胞反應。
Cell-mediated immunity can be evaluated by either radioactive or non-radioactive methods. Despite the availability of current non-radioactive assays, radioactive assay is still commonly employed due to its relatively high sensitivity and reliability. Nevertheless, major concerns have been raised about radiation hazard and safe disposal of radioactive waste. In this study, three non-radioactive assays were compared as to the sensitivity to measure the responsiveness of ICR mouse lymph node cells to soluble antigens extracted from CFW mouse sarcoma-180 whole cells, membranes and internal components. In cell-bead co-culture system, according to the degree and type of cellular accumulation, the reaction was recorded as cellular response index. Both amido black assay and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) based colorimetric assay were used for the spectrophotometric quantification of cell proliferation in response to stimulants. The results consistently showed that lymph node cells from S-180 immunized mice significantly responded to S-180 antigens but not to unrelated stimulant-bovine serum albumin. Among three S-180 antigens, whole cell antigen was apparently the best stimulant for S- 180 sensitized cells. As to sensitivity was concerned, MTT assay was the best one due to its ability to significantly differentiate the responsiveness to three S-180 antigens. The ability of cellular stimulation from high to low was in an order of whole cell antigen, internal antigen, and membrane antigen. Amido black assay could only marginally distinguish the responsiveness to S-180 membrane and internal antigens. In cell-bead co-culture system, cellular response index reached the highest level when immune cells were cultured with beads coated with S-180 whole cell antigen. However, no significant difference was observed when beads coated with internal or membrane antigen was used. In conclusion, nonradioactive methods are safe and also relatively sensit
Other Identifiers: AFD4935D-760C-5854-2615-CFB74B53ABE1
Appears in Collections:生物學報

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