产品说明书
FAQ
COA
已发表文献
产品描述
碱性磷酸酶(Alkaline Phosphatase, AP/ALP/AKP/ALKP/ALPase/Alk Phos)是一类水解酶，其通过水解磷酸单酯的作用将底物分子上的磷酸基团除去掉并生成磷酸根离子和自由的羟基。在碱性条件下以核苷酸、蛋白质和生物碱等作为去磷酸化作用的底物。该酶是一组同功酶的统称，如小牛肠碱性磷酸酶(Calf Intestinal AlkalinePhosphatase, CIAP/CIP)被广泛用于抗体标记并最终用于蛋白和核酸的检测等，也常用于DNA或RNA 5’和3’末端的去磷酸化(去单磷酸化)等。
BCIP/NBT是碱性磷酸酶的常用底物，在碱性磷酸酶的作用下BCIP会被水解，水解产物会和NBT发生反应形成不溶性的深蓝色至蓝紫色的NBT-formazan物质。
本试剂盒可用于细胞或组织的碱性磷酸酶显色，包括诱导多功能干细胞iPS的鉴定，也可以用于结合有碱性磷酸酶的膜的Western显色检测等。
产品组分
|
组分编号 |
组分名称 |
包装详情 |
|
40749-A |
碱性磷酸酶染色缓冲液 |
100 mL |
|
40749-B |
碱性磷酸酶染色液BCIP（300×） |
350 µL |
|
40749-C |
碱性磷酸酶染色液NBT（150×） |
700 µL |
运输和保存方法
冰袋运输。4℃保存，有效期1年，染色液BCIP和染色液NBT需避光保藏。
注意事项
1）本产品仅限科学研究。
2）BCIP对人体有刺激性，NBT对人体有害，请注意适当防护。
3）为了您的安全和健康，请穿实验服并戴一次性手套操作。
4）本产品仅作科研用途！
染色步骤
1、用合适的洗涤液对于组织切片或细胞样品或膜在与碱性磷酸酶标记的抗体或探针孵育后，洗涤3-5次，每次3-5 min。对于检测内源性碱性磷酸酶的组织或细胞样品，在固定后，也用合适的洗涤液洗涤3-5次，每次3-5 min。
2、按照 缓冲液：染色液BCIP：染色液NBT=3 mL: 10 µL:20 µL的比例配制适量体积的AP染色工作液。
3、在最后一次洗涤后去除洗涤液，加入适量AP染色工作液。
4、室温避光孵育5-30 min或更长时间(最长达24 h)，直至显色达到预期深浅效果。
5、去除AP染色工作液，用蒸馏水洗涤1-2次即可终止显色反应。
6、对于组织切片或细胞样品，在显色反应终止后如有必要可用中性红染色液(neutral red staining solution)染色观察。对于膜样品，显色反应终止后可至室温晾干避光保存。
Q: 使用碱性磷酸酶染色试剂盒（40749ES60），细胞，本身就是高表达碱性磷酸酶，做了处理之后可能就稍微低表达一些，拍出来的图，是细胞一整个轮廓是染色的？
A: 颜色深浅可以反应碱性磷酸酶的多少，但是微量差别可能肉眼无法观察，可以进行颜色灰度值测定（此方法可能会存在结果不稳定问题），可参考 WB 条带的计算方法。
Q:这个可以染胚胎干细胞吗？
A: 根据说明书中是可以的。本试剂盒可用于细胞或组织的碱性磷酸酶显色，包括诱导多功能干细胞iPS 的鉴定，也可以用于结合有碱性磷酸酶的膜的 Western 显色检测等。
Q:这个可以用于判断细胞活性不？
A: 不能用于细胞活性检测，本试剂盒主要用于细胞或组织的碱性磷酸酯酶显色包括诱导多功能干细胞 iPS 的鉴定，也可以用于 Western 等结合有碱性磷酸酯酶的膜的显色检测。同时也可以用于细胞或组织内源性的碱性磷酸酯酶显色。 检测细胞活性老师可以用cck-8 等试剂盒。
Q: 使用 24 孔板做成骨细胞分化后的碱性磷酸酶染色，请问可以使用本产品吗？
A: 可以的。
Q: 配好的碱性磷酸酶工作液可以重复是使用么，工作液如何保存，保存时间多久？
A: BCIP/NBT 染色工作液需现配先用，不能重复使用。
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