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[其它临床检测] 求:肺泡换洗液细胞分类的流式检测方法

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发表于 2012-5-30 17:09:02 | 显示全部楼层 |阅读模式
悬赏1流星已解决
临床要开展肺泡换洗液标本的细胞分类检测,
各位老师和同仁如果有该方面的方法和知道意见建议,请您教我。
1.标本如何处理?
2.换洗液细胞分类如何设计?

不胜感激

最佳答案

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效果还可以,排除个别老鼠取样问题,不同组的Treg百分比基本和实验预期一致. 如下图,是在CD4+细胞中分析,纵坐标是CD25,连续表达;横坐标是FOXP3.
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发表于 2012-5-30 17:09:03 | 显示全部楼层
效果还可以,排除个别老鼠取样问题,不同组的Treg百分比基本和实验预期一致.
如下图,是在CD4+细胞中分析,纵坐标是CD25,连续表达;横坐标是FOXP3.
11.jpg
流式中文网FlowGuard®流式专用保存液,无需冻存,稳定保护各类流式样本,从容完成实验
发表于 2012-5-30 20:49:15 | 显示全部楼层
是肺泡灌洗液吧?
听说过,但是没做过,帮你找篇参考文献,他们是做辅助性t细胞,里面有灌洗需要的液体,处理步骤,看看是否够?

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3127771/?tool=pubmed
Bronchoalveolar lavage (BAL) was performed by infusing three aliquots of 60 ml of sterile sodium chloride (NaCl), pH 7.3 at 37°C that were gently sucked back after each infusion and pooled into a container placed in iced water. The recovered fluid was immediately transported to the laboratory for analysis.

Flow cytometry analysis
BAL -lymphocyte subsets were determined using flow cytometry. BAL cells were centrifuged and diluted to a final concentration of 106 cells/ml. For each test, 10 μl of antibody solution was added to 200 μl of cell suspension and allowed to bind for 30 minutes at 4°C in darkness. Red blood cells were lysed with 2 ml FACS™ Lysing solution (Becton Dickinson Immunocytometry Systems, San Jose, CA, USA) for 10 minutes at room temperature. The remaining cells were then washed by adding PBS to the tubes and centrifuged at 4°C for 10 minutes, 300 g and repeated once. Cells were thereafter fixed with 500 μl CellFIX™ (Becton Dickinson Immunocytometry Systems, San Jose, CA, USA) before analysis using a FACSCalibur™ (Becton Dickinson) flow cytometer. The lymphocyte population was gated on their physical characteristics in a region according to their characteristic forward scatter (FCS) and side scatter (SSC) profiles, as previously reported [6]. 3,000 total events were collected in CD3+ gate per sample

流式中文网FlowGuard®流式专用保存液,无需冻存,稳定保护各类流式样本,从容完成实验
发表于 2012-5-31 10:20:19 | 显示全部楼层
这篇文章是2001年的,供参考:http://www.flowcyto.cn/bbs/forum.php?mod=viewthread&tid=1571
流式中文网FlowGuard®流式专用保存液,无需冻存,稳定保护各类流式样本,从容完成实验
发表于 2013-8-26 10:54:32 | 显示全部楼层
倪老师提供了好多好文章!我们这边最近也在开展这项目,还在摸索中,流式只看淋巴细胞中CD4/8/3的情况,灌洗液中细胞分类是制成涂片镜下观察计数
组织样本处理不好?流式中文网原研的魔滤®魔杵®套装,低成本解决,高质量收获
发表于 2013-8-26 11:09:32 | 显示全部楼层

成功了分享一下经验吧:)
流式中文网FlowGuard®流式专用保存液,无需冻存,稳定保护各类流式样本,从容完成实验
发表于 2013-8-27 09:29:13 | 显示全部楼层
使用流式检测肺泡灌洗液,想比较镜检要好很多,根据CD4/8/3的上升下降,可以很多肺部疾病的情况。
流式中文网FlowGuard®流式专用保存液,无需冻存,稳定保护各类流式样本,从容完成实验
发表于 2013-8-27 11:00:08 | 显示全部楼层
绿茶or红茶 发表于 2013-8-27 09:29
使用流式检测肺泡灌洗液,想比较镜检要好很多,根据CD4/8/3的上升下降,可以很多肺部疾病的情况。 ...

能举几个例子么?比如升高到多少,判断为什么疾病可能?
组织样本处理不好?流式中文网原研的魔滤®魔杵®套装,低成本解决,高质量收获
发表于 2013-9-2 22:27:56 | 显示全部楼层
做过大鼠的肺泡灌洗液的Treg流式检测,检测Treg markerCD4,CD25,FOXP3.冲洗肺泡,收细胞,表染,固定破膜(FOXP3专用),染FOXP3,上机。通过检测Treg的百分比来和不同药物处理的group关联分析。
组织样本处理不好?流式中文网原研的魔滤®魔杵®套装,低成本解决,高质量收获
发表于 2013-9-2 22:45:33 | 显示全部楼层
judy006 发表于 2013-9-2 22:27
做过大鼠的肺泡灌洗液的Treg流式检测,检测Treg markerCD4,CD25,FOXP3.冲洗肺泡,收细胞,表染,固定破膜 ...

效果如何?分群清楚么?
组织样本处理不好?流式中文网原研的魔滤®魔杵®套装,低成本解决,高质量收获
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