TopoIIBufferissuppliedasa10xStocksolutionintwopartsthatmustbemixedpriortouse:
- 10XIncompleteTopoIIAssayBufferAcontainsthefollowing:0.5MTris-HCl(pH8.0),1.5MNaCl,0.1MMgCl2,5mMdithiothreitol.
- 10XATPBufferBcontains20mMATPinwater.
Thesetwobuffersmustbemixedtoprepareafinal5Xworkingstockofassaybuffer. FurThermore,thefinal5XCompleteBuffercontainsbothAandBandshouldbemadefreshforeachexperiment. Donotfreezethe5XCompleteBuffer;itisstablefor24hoursonice. ToPreparethe5XCompleteBuffer,mixequalvolumesofAandB(0.1mlofAand0.1mlofB)andplaceat4°Coronice.
QualityControlAnalysis:
The5XCompleteTopoIIAssayBuffer(containingequalpartsAandBabove)isoptimizedandtestedfordecatenationandrelaxationactivityofeukaryotictopoII. TheCompleteBuffershouldbemadefreshforeach
experiment.