Luc-Screen™ 延长辉光荧光素酶报告基因检测系统

For Fetal Bovine Serum (FBS), there is no difference between 10% and 5% FBS. It is worthwhile to note that the type of serum does affect the luciferase assay. We tested the effect of 6 different types of sera (FBS, heat-inactivated FBS, dialyzed FBS, charcoal-treated FBS, donor adult bovine serum, and donor equine serum) on secreted luciferases, e.g. Gaussia, Gaussia-Dura, and Cypridina Luciferases. No noticeable difference was observed among different types of sera except for donor adult bovine serum. Unknown factors in donor adult bovine serum caused up to 35% inhibition in secreted luciferase activity. Therefore, we don’t recommend using donor adult bovine serum for Gaussia, Gaussia-Dura, and Cypridina Luciferases.

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The problems with white plates are related to cross-talk blocking, surface reflectivity and tendency to phosphorescence. We do not recommend using white plates because they give much higher background even if adapted to darkness for 10 minutes. The white plastic does not block the light completely, and some portion of the signal will go through the plastic and will be seen when you measure adjacent wells. The typical situation is if you change from black to white plates, where the background is about doubled and the signal is increased about 10 times. Black plates are recommend for best signal to noise ratio even though the RLU values will be lower. 

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Gaussia, Gaussia-Dura, and Cypridina Luciferase are stable in the culture media for greater than 24 hours. Generally, after transfection of CMV driven constructs, we see ever increasing secreted signal between 24 and 72 hours.

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The earliest detectable signal will depend on the strength of the promoter. In general, with a strong promoter (such as CMV), significant signal over background can be seen in as little as 20 minutes. However, with a weaker inducible promoter, significant signal over background may take 1-2 hours after induction.

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We always recommend the luciferases to be read without filters for single luciferase assays. Use of filters reduces the amount of signal captured that may lead to decrease in sensitivity. Filters are required only for dual-spectral assays.

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