Firefly includes in each well a positive control called “X-control”, to ensure that the reactions occurred robustly and consistently in each well, and a negative control “Blank” to control for background fluorescence. In addition, we recommend that you choose endogenous controls, microRNAs known not to fluctuate in your system, to allow for normalization between sample types and treatments. Firefly suggests the following endogenous controls for most samples but you should confirm that these work for your samples:
- Human: RNU44, RNU48, RNU6B
- Mouse: snoRNA202, snoRNA234
- Cell Lines: miR-16-5p (same sequence for human, mouse, and rat)
- C. elegans: U18
For more detailed explanations see Controls.
Note: Serum and plasma do not contain RNUs so we recommend miR-16, miR-451a, and mir-486 instead.