碧云天生物技术-CFDA SE细胞增殖与示踪检测试剂盒(C0051)

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CFDA SE细胞增殖与示踪检测试剂盒

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产品编号: C0051

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产品包装:2000次

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产品编号	产品名称	产品包装	产品价格
C0051	CFDA SE细胞增殖与示踪检测试剂盒	2000次	1001.00元

CFDA SE细胞增殖与示踪检测试剂盒(CFDA SE Cell Proliferation Assay and Tracking Kit)是碧云天研发的基于一种新型荧光探针CFDA SE的用于细胞增殖检测和细胞荧光示踪的检测试剂盒。CFDA SE目前被广泛用于替代MTT法或[³H]-thymidine掺入等其它细胞增殖检测方法。
基于CFDA SE荧光标记的细胞增殖检测和[³H]-thymidine掺入、BrdU标记获得的检测结果完全一致，但同时可以提供更多的细胞增殖信息。使用CFDA SE检测可以提供整个细胞群中有多少比例的细胞分裂了1次、2次或更多次数，同时如果和其它荧光探针联用，可以获取不同分裂次数细胞的其它相关信息。
CFDA-SE的全称为Carboxyfluorescein diacetate, succinimidyl ester，分子式为C₂₉H₁₉NO₁₁，分子量为557.47，CAS
number为150347-59-4。CFDA SE可以通透细胞膜，进入细胞后可以被细胞内的酯酶(esterase)催化分解成CFSE，CFSE可以偶发性地(spontaneously)并不可逆地和细胞内蛋白的Lysine残基或其它氨基发生结合反应，并标记这些蛋白。在加入荧光探针CFDA SE后大约24小时，即可充分标记细胞。被CFDA SE标记的非分裂细胞的荧光非常稳定，稳定标记的时间可达数个月。CFDA SE标记细胞的荧光非常均一，比以前使用的其它细胞示踪荧光探针例如PKH26的荧光更加均一，并且分裂后的子代细胞的荧光分配也更均匀。
由于CFDA SE标记细胞的荧光非常均匀和稳定，每分裂一次子代细胞的荧光会减弱一半，这样通过流式细胞仪检测就可以检测出没有分裂的细胞，分裂一次的细胞(1/2的荧光强度)，分离两次的细胞(1/4的荧光强度)，分裂三次的细胞(1/8的荧光强度)以及类似的其它分裂次数的细胞。采用CFDA SE通过流式细胞仪检测获得的检测结果参考右图。每一个峰代表一种分裂次数的细胞，从右至左的峰通常依次为分裂0次、1次、2次、3次等次数的细胞。分裂次数较多后，分裂0次或1次等没有分裂或分裂次数较少的细胞会逐渐减少直至检测不到。
使用CFDA SE可以检测分裂多达8次或更多次数的细胞增殖。
目前CFDA SE标记细胞后通常用流式细胞仪进行细胞增殖检测。最常用于淋巴细胞的增殖检测，也可以用于成纤维细胞、NK细胞等其它细胞的增殖检测，甚至还可以用于细菌增殖的检测。
CFDA SE标记细胞呈绿色荧光，检测时的激发波长可以选择488nm，此时的发射波长为518nm，使用流式细胞仪检测时可以采用FL1 detection channel。CFDA SE标记的细胞也可以用荧光显微镜进行观察。
CFDA SE标记的细胞无论在体外还是体内都不会使邻近细胞染色。即CFDA SE荧光探针完成标记后不会从一个细胞转移到邻近细胞。
CFDA SE标记细胞仅需5-15分钟即可完成。对于不同细胞，最佳标记时间需自行摸索。
本试剂盒提供了配制CFDA SE所需的溶液以及CFDA SE进行细胞标记时所需的配套试剂。
如果每个检测样品的荧光探针标记体积为1ml，本试剂盒共可以检测2000个样品。

包装清单：

产品编号	产品名称	包装
C0051-1	CFDA SE	1管
C0051-2	CFDA SE溶剂	1ml/管，共两管
C0051-3	CFDA SE细胞标记液(10X)	100ml/瓶，共两瓶
—	说明书	1份

保存条件：
-20℃保存，半年有效。其中CFDA SE需避光保存。
注意事项：
CFDA SE配制成储存液后宜在一个月内使用完毕，最长不宜超过2个月。CFDA SE易被水解，在水溶液中会很快变质。请在使用过程中避免接触水。在标记细胞的过程中和水接触是在许可的范围内的。
CFDA SE溶剂在4℃、冰浴等较低温度情况下会凝固而粘在离心管管底、管壁或管盖内，可以20-25℃水浴温育片刻至全部融解后使用。
荧光染料均存在淬灭问题，请尽量注意避光，以减缓荧光淬灭。
本产品仅限于专业人员的科学研究用，不得用于临床诊断或治疗，不得用于食品或药品，不得存放于普通住宅内。
为了您的安全和健康，请穿实验服并戴一次性手套操作。

使用说明：
1. 检测前的准备：
a. CFDA SE储存液(1000X)的配制：用试剂盒中所带的共2ml CFDA SE溶剂溶解CFDA SE，即可配制成CFDA SE储存液(1000X)。配制完成后分装到试剂盒所提供的原来装CFDA SE溶剂的管子中，并做好标记。具体的操作步骤如下：取1ml CFDA-SE溶剂加入到CFDA SE中，充分溶解后取0.5ml至原1ml CFDA-SE溶剂管子中，再从另外一管CFDA SE溶剂中取0.5ml至该管中并混匀；取另外的0.5ml CFDA SE至另外一管0.5ml的CFDA SE溶剂管中并混匀。配制好的两管CFDA SE储存液均为1000X，-20℃避光保存，最好能-70℃避光保存。-20℃避光保存宜在一个月内使用完毕，最长不宜超过2个月。-70℃避光保存可以适当延长使用时间。
b. CFDA SE细胞标记液的配制：准备适量无菌的细胞培养级纯水，根据实验需要配制适量的CFDA SE细胞标记液。例如，取20ml CFDA SE细胞标记液(10X)，加入180ml细胞培养级纯水，混匀后即为CFDA SE细胞标记液。配制好的CFDA SE细胞标记液可以4℃保存，长时间不用可以-20℃保存。
2. 标记和检测：
a. 用1ml CFDA SE细胞标记液悬浮100万至500万细胞，置于15ml离心管内。
b. 用CFDA SE细胞标记液稀释CFDA SE储存液(1000X)至2X。例如取2微升CFDA SE储存液(1000X)至1ml CFDA SE细胞标记液中，混匀后即为CFDA SE储存液(2X)。
c. 把1ml CFDA SE储存液(2X)加入到步骤2.a中含有1ml待标记细胞的15ml离心管内，轻轻混匀。
d. 37℃孵育10分钟。
e. 立即在15ml离心管内加入约10ml完全细胞培养液(含血清)，室温颠倒数下混匀。
f. 室温离心去上清，再用5-10ml完全细胞培养液洗涤一次。
g. 再加入5-10ml完全细胞培养液，37℃孵育5分钟，以促进CFDA SE在细胞内的驻留及未反应的CFDA SE进入完全细胞培养液。离心去上清，完成最后一次洗涤。
h. 随后即可按照细胞的正常培养方法进行培养。可以在荧光显微镜下直接观察标记效果，也可以在培养适当时间后用流式细胞仪检测细胞增殖，或用于特定目的的细胞示踪。标记的细胞也可以用于活体动物的移植，并用荧光进行示踪。标记的细胞呈绿色荧光。

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