Notch1 receptor drug validation assay

[haquee3]

I am trying to screen around 600 different compounds to see if any will have antagonist properties on the human notch1 receptor, which is permenantly activated resulting in a tumor.

I will be adding the max concentrations of the different compounds to the cancer tumor cells, all done on 96 well plates.

The problem is how to measure the response from the different compounds. I believe the notch receptors, when activated, can be measured by gene expression. However i can not seem to find a suitable assay that will show me which compounds has inhibited the receptor.

I am willing to spend around £700 for the assay.

If anyone has any suggestions they will be much appreciated, thank you.

 

[CharonY]

Not sure what you are asking. Are do you want a way to measure inhibition or one that allows you to identify the inhibiting compound from a mixture. The latter is difficult and I guess not what you are asking. For the former typically a luminescence (or fluorescence) reporter is often used coupled to a Notch pathway responsive reporter. Though technically you could also just conduct qPCR on responsive genes, but for the number of compounds it may not be the most efficient approach.

[hypervalent_iodine]

Unless I have misunderstood, the OP is looking to perform a high throughout screening assay. A quick Google search suggests this has been done before, so you should be able to find a suitable protocol for identifying compound hits easily enough. See here, for example. 

[CharonY]

Well, most of these assays are  reporter-based.

[haquee3]

I believe that a luminescence assay, that was suggested, may be the most suitable. Just need to find an appropriate kit to buy.

Anyways, thank you for the help guys.

[CharonY]

One quick tip for these assays, often you can save a bit of time if you multiplex at the beginning to pick out suitable candidates. May not work if the compounds have cross-reactivity, though.