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当前位置: 首页 > 文章 > 肉仔鸡肠道B淋巴细胞Ca~(2+)浓度流式细胞术测定方法的建立 西北农林科技大学学报(自然科学版) 2011,39 (3) 13-16
Position: Home > Articles > Establishment of the method to detect Ca~(2+) of intestinal B lymphocytes of broiler chickens with flow cytometry Journal of Northwest A & F University(Natural Science Edition) 2011,39 (3) 13-16

肉仔鸡肠道B淋巴细胞Ca~(2+)浓度流式细胞术测定方法的建立

作  者：
王益兵;姚军虎;覃定奎;李朝云;杨小军
单  位：
西北农林科技大学动物科技学院
关键词：
流式细胞术;二十碳五烯酸(EPA);B淋巴细胞;Ca2+浓度
摘  要：
【目的】建立以流式细胞术结合Fluo-3-AM测定肉仔鸡肠道B淋巴细胞胞内Ca2+浓度的方法。【方法】分离培养21 d肉仔鸡肠道B淋巴细胞,以不同质量浓度(0,5,10,15,20μg/mL)脂多糖(LPS)刺激B淋巴细胞,测定B淋巴细胞刺激指数(SI),确定B淋巴细胞增殖的最适LPS质量浓度,建立肉仔鸡B淋巴细胞体外增殖试验模型。以最佳质量浓度LPS刺激培养72 h的肉仔鸡肠道B淋巴细胞为研究对象,应用Fluo-3-AM作为钙指示剂,建立肉仔鸡肠道B淋巴细胞胞内Ca2+浓度的流式细胞术测定方法,并用该测定方法探讨不同质量浓度(0,20,40,60,80μg/mL)二十碳五烯酸(EPA)对肉仔鸡肠道B淋巴细胞胞内的Ca2+浓度的影响。【结果】LPS的最佳质量浓度为10μg/mL。建立了以Fluo-3-AM为标记的肉仔鸡肠道B淋巴细胞胞内Ca2+浓度流式细胞仪测定方法。EPA可极显著降低B淋巴细胞胞内Ca2+浓度(P<0.01)。【结论】Fluo-3-AM结合流式细胞术可检测肉仔鸡肠道B淋巴细胞胞内的Ca2+浓度,且EPA极显著降低了B淋巴细胞胞内的Ca2+浓度。
译  名：
Establishment of the method to detect Ca~(2+) of intestinal B lymphocytes of broiler chickens with flow cytometry
作  者：
WANG Yi-bing,YAO Jun-hu,QIN Ding-kui,LI Chao-yun,YANG Xiao-jun (College of Animal Science and Technology,Northwest A&F University,Yangling,Shaanxi 712100,China)
关键词：
flow cytometry;EPA;B lymphocytes;Ca2+concentration
摘  要：
【Objective】 The study was done to establish a method of detecting intracellular Ca2+ in intestinal B lymphocytes of broiler chickens with Fluo-3-AM by flow cytometry.【Method】 Healthy 21 d broiler chickens were used for our experiments.The 3 cm jejunum was removed from 21 d broilers,the intestinal B lymphocytes were isolated and cultured in vitro.LPS was used at different concentrations(0,5,10,15,20 μg/mL) for stimulation of intestinal B cell cultures.The optimal conditions for LPS stimulation of in vitro B lymphocyte proliferation was determined by choosing the LPS concentration that gave the maximal stimulation index.Intestinal B lymphocytes were stimulated by LPS,while the system was added different concentrations of EPA(0,20,40,60,80 μg/mL),and then the cell was loaded with Fluo-3-AM as a calcium indicator.The intracellular Ca2+ in intestinal B lymphocytes of broilers was examined with Fluo-3-AM by flow cytometry.【Result】 The optimal concentration of LPS was 10 μg/mL.The fluorescence intensity of B lymphocytes with Fluo-3-AM was increased significantly.The method was established to detect the intracellular Ca2+ in intestinal B lymphocytes of broiler chickens.EPA can significantly reduce the Ca2+ of B lymphocytes of broilers(P<0.01).【Conclusion】 EPA significantly decreased the intracellular Ca2+ of B lymphocytes,and using Fluo-3-AM and flow cytometry could detect the intracellular Ca2+ of B lymphocyte.

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