四色多重荧光免疫组化染色试剂盒(鼠兔通用二抗)
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abs50012-100T | 1-2周 | ¥20250.00 | - + |
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- 实验方法 实验条件
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- 微波修复抗原每一轮染色都要做吗?可以用抗体洗脱液替代吗?抗体洗脱液用在哪个步骤?
- 染第一个抗体之前做抗原修复,是为了使被石蜡或者固定剂封闭的抗原决定簇重新暴露出来,使得抗体能够识别到抗原;在染后面每一个抗体之前的抗原修复,目的是为了去除掉上一轮染色的抗体和残留没有结合上的染料,其实更准确的叫法是“抗体洗脱”,也就是说第一次的抗原修复是真正的抗原修复,后面的抗原修复实际上是“抗体洗脱”,“抗体洗脱”可以用抗体洗脱液(abs994)替换,特别是细胞、冰冻切片或者骨样本,但第一次抗原修复不能被抗体洗脱液替代,冰冻切片可以不进行抗原修复,染色效果欠佳时,可以考虑尝试进行抗原修复,推荐冰冻切片快速抗原修复液(5×)(abs9207)。
- 脱蜡水合步骤中“10%中性福尔马林浸片10min,灭菌水洗片1min,重复3次”目的是什么?我们以前做石蜡切片的时候没有这一步。
- 这是后固定的步骤,固定充分的组织可以省略此步。
- 做好的多色片子,无法进行及时扫描,该如何保存?
- 需要避光防震,在4℃条件下保存
- 有详细的实验操作流程吗?
- 有的,我们有完整的操作视频,您可以点击链接观看完整视频。
- 样本可以做成冰冻切片吗?
- 可以的。抗原修复步骤不推荐用加热的方法,推荐用抗体洗脱液(abs994)
- 所有操作要避光吗?
- 不需要,在常规环境下操作就可以,染料的荧光强度很高。
- 抗体修复液每一轮修复都要更换吗?
- 是的,每一轮都要根据抗体选择相应的修复液。
- 染料的选择会影响后染色吗,有共定位的话怎么办?
- 共定位的情况,我们可以通过单一通道观察来更好的检测各指标的分布情况,染料不会对染色有影响,串色的情况需要预实验做充足,包括抗体稀释比例,染料与放大液比例,修复条件的确定。
- 多色用的一二抗各是什么种属?
- 多色的优势在于一二抗种属无需特别要求,一抗根据样本种属选择既可,二抗根据一抗选择即可,多个抗体组合,种属不会有影响。
- 抗体洗脱和微波修复哪种方式洗脱效果更好?
- 微波修复洗脱更稳定,抗体洗脱液对大多数抗体都管用,难洗的抗体延长洗脱时间就可以啦!
- 实验用时真的一天能做完吗?
- 相比普通的组织荧光实验,我们一抗的孵育时间只需室温30min,二抗和信号放大步骤只需孵育10min,能大大节约时间,五色左右可在一天内做完。
- 多色样本的寄送有条件限制吗?
- 正常的石蜡切片和组织芯片很稳定,只要防震盒常温运输即可,冰冻切片的话,需要干冰低温运输。
- 我的组织还有GFP蛋白,给结果的时候能排除干扰吗?
可以的,结果分析的时候可以避免绿色通道,分析软件赋予的颜色是伪彩色,可以更换其他颜色。
- 我这边提供抗体的话,抗体需要满足什么条件吗?
抗体需要要满足IHC的实验应用,各品牌供应商都会提供相应的实验验证的,爱必信组化抗体经过严格实验验证,是不错的选择。
- 如果我能提供抗体给你们做服务的话,需要的抗体量是多少?
需要的抗体量要根据样本数来,但一般来讲,50-100ul的抗体足够完成预实验+正式实验了。
- 在指标与染料的搭配以及染色顺序上,有什么规律吗?
- 并没有一个特别的线性关系,和蛋白的种类、表达量以及修复条件都有关系,所以会多因素考虑,多尝试,预实验很重要。
- TSA技术原理中有抗体洗脱的步骤,但是在你们的试剂盒步骤说明中却没有?
试剂盒步骤中的抗原修复过程就是抗体洗脱过程。
- B细胞、T细胞、DC细胞常用maker能推荐下吗?
T细胞:CD3、CD4、CD8、FoxP3(再细化还有:GranzymeB、PD-1、TIM-3、CD45)
B细胞:CD19 ;
DC细胞:CD8α、CD11b
- 荧光染料激发光和发射光波长分别是多少?
染料名称
保存温度
波长
相似荧光染料
激发波长
发射波长
DAPI
-20℃
360
461
DAPI
TSA单色荧光染料480
430
480
Opal480
TSA单色荧光染料520
490
520
FITC/AF488/Opal520
TSA单色荧光染料540
520
540
AF514/Opal540
TSA单色荧光染料570
550
570
CY3/AF555/Opal570
TSA单色荧光染料620
590
620
AF594/Opal620
TSA单色荧光染料650
640
660
AF610/Opal650
TSA单色荧光染料700
682
702
AF680/CY5.5/Opal690
TSA单色荧光染料770
740
780
Opal780
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