Name: preparing t cell growth factor from rat splenocytes
Uploaded: 2007-10-31T22:24:33
Description: Watch this Scientific Journal Video about Preparing T Cell Growth Factor from Rat Splenocytes at JoVE.com

<ol><li>以Lewis大鼠脾脏（160 - 200G之间的老鼠是最好的）。在培养皿菜含有PBS +抗生素（PBS - PS），使用细胞过滤器Dilacerate冰。放入50毫升管。填写用PBS - PS。 </li><li>自旋为10分钟，在4 ° C至沉淀细胞。 </li><li>清洗细胞两次。 </li><li>悬浮颗粒在NH 4氯0.15米（5毫升每脾）。混合3分钟就冰吸管轻轻地，不断溶解红细胞。填充介质管。 </li><li>自旋为10分钟，在4 ° C至沉淀细胞。 </li><li>清洗细胞两次。 </...

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	<li>Beeton, C., Barbaria, J., Devaux, J., Benoliel, A. -. M., Gola, M., Sabatier, J. -. M., Bernard, D., Crest, M., Beraud, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selective+blocking+of+voltage-gated+K++channels+treats+experimental+autoimmune+encephalomyelitis+and+inhibits+T-cell+activation.">Selective blocking of voltage-gated K+ channels treats experimental autoimmune encephalomyelitis and inhibits T-cell activation.</a> J. Immunol. 166, 936-944 (2001).</li><li>Beeton, C., Wulff, H., Barbaria, J., Clot-Faybesse, O., Pennington, M., Bernard, D., Cahalan, M. D., Chandy, K. G., Beraud, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selective+blockade+of+T+lymphocyte+K++channels+ameliorates+experimental+autoimmune+encephalomyelitis,+a+model+for+multiple+sclerosis.">Selective blockade of T lymphocyte K+ channels ameliorates experimental autoimmune encephalomyelitis, a model for multiple sclerosis.</a> Proc. Natl. Acad. 98, 13942-13947 (2001).</li><li>Mor, F., Cohen, I. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Propagation+of+Lewis+rat+encephalitogenic+T+cell+lines:+T-cell-growth-factor+is+superior+to+recombinant+IL-2.">Propagation of Lewis rat encephalitogenic T cell lines: T-cell-growth-factor is superior to recombinant IL-2.</a> J. Neuroimmunol. 123, 76-82 (2002).</li></ol>

我们准备从Lewis大鼠脾细胞TCGF，因为我们经常从天真这株收获血清和thymi Lewis大鼠T细胞系在体外刺激大鼠安乐死。这TCGF，可以被用来促进从其他的大鼠株T细胞系的生长和生存。 TCGF也可准备从其他菌株的大鼠。

为了准备完全培养液中添加以下的RPMI 1640培养液和无菌过滤瓶500毫升：10％FCS的1瓶，PSG; 5毫升的RPMI维生素;丙酮酸钠5毫升，5毫升的非必需氨基酸; 50 UM β-巯基乙醇; 2微克/毫升伴刀豆球蛋白A。

preparing t cell growth factor from rat splenocytes

抗原特异性T细胞株或文化克隆的维修需要轮抗原诱导激活细胞扩张 1,2阶段分开。白细胞介素2除了在扩张阶段的文化传媒，是必要措施，防止细胞死亡，并足以维持短期的T细胞系，但已被证明能增加Th1细胞极化3。 T细胞生长因子（TCGF），其中包含混合的细胞因子白细胞介素2的更换是更有效地保持长期的T细胞系在体外培养3比白细胞介素2。此外，TCGF可以很容易地在实验室中大量的准备和重组白细胞介素2比便宜得多。 在这里，我们展示如何准备从大鼠脾细胞培养上清TCGF。在此过程中，我们收获脾脏从幼稚刘易斯安乐死大鼠胸腺和血液采集。我们准备从脾单细胞悬液，析渗透压冲击的红血细胞，而种子在培养基中的脾。与非选择性地激活所有大鼠T淋巴细胞，诱导细胞因子的产生，有丝分裂原刀豆蛋白A的细胞受到刺激。文化supernantant收集48小时后andexcess伴刀豆球蛋白A是必然的α甲基mannoside，以防止它激活T细胞，TCGF将增加线路。 TCGF是无菌过滤，分装，储存于-20 ° C

Watch this Scientific Journal Video about Preparing T Cell Growth Factor from Rat Splenocytes at JoVE.com

Preparing T Cell Growth Factor from Rat Splenocytes

我们描述的T细胞生长因子在体外扩增抗原特异性大鼠T淋巴细胞线的准备。

准备从大鼠脾细胞T细胞生长因子

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