| 摘要: |
| 【目的】探讨磺酰罗丹明B法(Sulforhodamine B,SRB)检测鸡外周血T淋巴细胞增殖反应的最佳条件。【方法】体外培养鸡外周血T淋巴细胞，选用SRB法对淋巴细胞含量(1×105，1×106，2.5×106，5×106，7.5×106和1×107 mL-1)、ConA质量浓度(5.0，7.5，10.0，12.5和15.0 μg/mL)、培养时间(36，48，60 h)3个因素进行组合设计培养T淋巴细胞，每个组合设3个重复，SRB显色后用酶标仪在492 nm波长处测量OD492值，计算刺激指数(Stimulation index，SI)，筛选SRB法检测鸡外周血T淋巴细胞增殖的最佳条件。【结果】SRB法检测鸡外周血T淋巴细胞增殖试验的最佳条件为：淋巴细胞含量1×106 mL－1},ConA质量浓度5.0 μg/mL,培养时间48 h。【结论】确定了体外培养鸡外周血T淋巴细胞增殖的最佳ConA质量浓度、培养时间及淋巴细胞含量，证实用SRB法检测鸡外周血T淋巴细胞增殖结果可靠，是切实可行的淋巴细胞数量与活性检测方法。 |
| 关键词: 鸡 外周血T淋巴细胞 细胞增殖 磺酰罗丹明B法 |
| DOI： |
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| 基金项目:农业部“948”引进项目(2009-Z45(2)) |
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| Screening the optimum SRB assay for chicken peripheral T-lymphocytes proliferation |
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| Abstract: |
| 【Objective】The optimal experiment conditions for SRB assay in chicken peripheral T-lymphocyte proliferation were investigated.【Method】The technology of cell in vitro culture to the peripheral T-lymphocyte of chicken was used.Three major factors which affect the proliferation,including the concentration of cell (1×105,1×106,2.5×106,5×106,7.5×106,and 1×107),the ConA density (5.0,7.5,10.0,12.5 and 15.0 μg/mL),and the cell culture time (36,48 and 60 h),were studied.The SRB assay and micro-plate reader with a wavelength of 492 nm were also performed.Meanwhile,the stimulation index was calculated.【Result】The optimal selection of SRB was:culture for 48 h,5.0 μg/mL ConA and 1×106 mL-1 cell concentration.【Conclusion】The determinate concentration of cell,ConA density and cell culture time were obtained.It shows that SRB assay is a reliable practical method to analyze the number of lymphocyte and detect their activity. |
| Key words: chicken peripheral T-lymphocyte cell proliferation SRB assay |
