Great EscAPeTM SEAP Fluorescence Detection Kit

Description

The Great EscAPeSEAP Fluorescence Detection Kit is a complete system for the detection of secreted alkaline phosphatase SEAP activity in conditioned media from cultures of mammalian cells expressing this enzyme. The kit contains reagents necessary for the detection of this enzyme activity using the fluorescent substrate 4-methylumbelliferyl phosphate MUP . Our products are to be used forResearch Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval. Back SEAP activity is highly specific, with a >20-fold increase in activity.HEK 293 cells were transiently transfected with a promoter construct containing CRE driving the expression of SEAP, or mock-transfected. 12 hr posttransfection, the cell culture media was replaced by media containing either 10 muM forskolin or plain cell culture media and incubated for 7 hr. Forskolin causes an increase in the level of cytosolic cAMP, which in turn activates CRE, driving the expression of SEAP, which is detectable in the media culture supernatant. The samples were assayed using the Great EscAPe SEAP Chemiluminescence Kit 2.0 and its protocol and analyzed on a BD Monolight 3096 Luminometer. Back Comparison of the sensitivities of SEAP and firefly luciferase.Parallel cultures of BHK cells were transiently transfected with the indicated amounts of either pSEAP2-Control Vector or a similar vector expressing luciferase. After 24 hr, SEAP activity was assayed in the appropriate culture media using the Great EscAPe chemiluminescent assay. Similarly, after 24 hr, cell lysates were prepared from the luciferase cultures, and luciferase activity was assayed using a commercial kit. RLU = relative light units. Back Back