发光分泌型碱性磷酸酶报告基因测定试剂盒 *发光剂 *Luminescent Secreted Alkaline Phosphatase Reporter Gene Assay Kit *Luminometric*详细如下: Luminescent Secreted Alkaline Phosphatase Reporter Gene Assay Kit 为胎盘碱性磷酸酶的分泌和膜结合形式提供了一种高度灵敏的检测方法。该试剂盒包含:化学发光底物、检测缓冲液、终止溶液、胎盘碱性磷酸酶标准品、与高通量筛选兼容的“混合和读取”检测方案。 分泌型碱性磷酸酶 (SEAP) 广泛用作报告基因,用于分析细胞培养物或动物中的基因表达、启动子和转录因子的活性。SEAP可以有效地分泌到培养基和血清中。SEAP 的这一独特特性使其便于对一段时间内的基因表达进行动力学分析。 Prepare placental alkaline phosphatase containing sample: Collect supernatants from transfected or control cells or prepare cell extracts if membrane-bound placental alkaline phosphatase was used for transfection (refer to Appendix for the preparation of cell extract). Note: The supernatant or cell extract can be stored at -70 ℃ for later use. Heat the culture supernatant or cell extract at 65°C for 10-30 min to inactivate endogenous non-specific alkaline phosphatase. Then cool down to room temperature. Chemiluminescent alkaline phosphatase substrate (Component A): Ready to use. Prepare alkaline phosphatase dilution buffer: Dilute 10X Assay buffer (Component B) to 1X with deionized water. Prepare dilutions of alkaline phosphatase: Dilute alkaline phosphatase standard (10 ug/mL - Component C) to 0.05 ug/mL (1:200) in dilution buffer. Then make fourfold serial dilutions to get the concentration of 12.5, 3.125, 0.78, 0.195, 0.05, and 0.012 ng/mL of alkaline phosphatase solution. Include a blank control. Note: Unused portion of diluted alkaline phosphatase solution should be discarded.