Certain compounds will quench the light emission in bioluminescent reactions.  Typically these are strong red dyes (however, the ViaLight ™ Plus Assay has been designed to function in all phenol red containing media commonly used in cell culture). 
Fetal Calf Serum also quenches but levels of FCS below 10% have no significant effect on assay sensitivity.  Concentrations above 10% may reduce the sensitivity of the assay.  When investigating dose-response relationships it is therefore strongly recommended that the test compounds are diluted in the complete media in which the cells are cultured. 
The quenching effect of an unknown compound can be tested by performing the assay in the presence of an ATP Standard (product code LT27-008).
The assay is based on the firefly luciferase enzyme which has its temperature optimum at 22°C, the test should, therefore, be performed at room temperature and all reagents should be allowed to equilibrate to this temperature before use.