Teaching Points for Enzyme Inhibition Assay


VDS Staff,

The assay will take a while and they may choose to do Run 2 another day.

Have them label 9 tubes A, B, C, D, E, F, G, H, I  for the samples – use the 1.7 ml micro centrifuge tubes

Then label 5 more 1.7ml tubes to put their stocks in.

500 mM Tris

50 mM Tris

MgCl2+

50%/50% water/DMSO

Enzyme

They should not be pipetting directly out of the Stock except to move it to their stock tube. This will prevent contamination of the stocks.

The will get a separate tube for pNPP – so they don’t need to make one for this

For the inhibitor compound, They need to choose which one of the 5 they want. They will get one tube of this from the White compounds box.

They also need to get one tube of the working solution of Orthovanadate.

After they have used the compounds – they should NOT put them back in the white box. If they are going to continue the 2nd assay on another day – they can save their compounds in a safe place where they can find them.

When they do the spectrophotometer – they should measure samples A –> I and then repeat the readings A–> I    (they will take the average of these two readings to get their Run 1 data).

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