I have 2 cell groups;
(WT)Wild-Type
(Subject)Wild-Type+Transfected Gene
I expose these cells to a particular drug at different doses.
I then look at calcium influx.
The drug is known (has been shown in many papers) to cause cell death and increase calcium influx.
I just assumed that the positive control would be the treated WT, and the negative control would be the untreated WT.
Do I really need a specific calcium influx inducer such as ionomycin or glutamate as a positive control for calcium assay/imaging, when the drug treatment itself is known to cause calcium influx?
If your goal is to analysed whether transfected cells are more or less sensitive to the drug than WT cells, you don't need a drug control.
However if you would like to verify if your drug is active, you need to challenge cell with the vehicle of your drugs versus your drug
I have no information about the construction of your transfected gene. However, a key question might be whether or not the procedure produced transfected cells. There may be some indicator available to check the efficiency and determine the proportion of transfected cells. So, check with the supplier of the gene construct.
Hi Martin,
The purpose of any control is to allow you to interpret the experimental output appropriately. So imagine a situation where the transfection reagent and/or the addition of DNA causes an increase in sensitivity to your drug - would you be able to identify this and separate it from an effect of the drug on the protein you have transfected? What are the other possible explanations that might give rise to the data you are seeing and how would you control for these in your experimental design?
If this is the same mode of action (calcium influx , e.g. by ionomycin), I would propose to additionally test this compound (comparatively) to get a "real" positive control .
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