The assay should be performed as outlined the assay protocol as this is how it has been validated.

This kit was developed to work in test tubes using the assay format outlined and we have not performed this assay in a 96 well microplate format. 

However, if you wish to do this in your own lab, the only part of the procedure that can possibly be performed in a microplate format is the GOPOD incubation and/or the absorbance read steps.

The 50 degree incubation of the GOPOD reagent with the sample could be done in the microplate format but the wells will need to sealed during the incubation. Obviously incubation cannot be done in a water bath but in an incubation oven. Also, the volumes need to be decreased by 10-fold i.e. 0.01 mL sample plus 0.3 mL GOPOD reagent. The lower volumes and potentially the heat transfer may introduce more error than observed in the higher volume glass test tubes. The glass test tubes in the water bath provide the best heat transfer and control to the reaction solution and I strongly recommend to use the test-tubes and then transfer to the microplate for reading if required.

Please note: When you decrease the volume of GOPOD Reagent you decrease the amount of glucose you can measure i.e. While using 3 mL of GOPOD Reagent you can measure up to 100 μg of glucose, if you decrease the volume of GOPOD Reagent 10-fold to 0.3 mL, you can only measure up to 10 μg of glucose. The same volumes would be required for your glucose standards and the calculation will change slightly.

Alternatively, if you really wish to use the microplate in this assay you can use the microplate as a means to read the absorbances rather than the entire incubation of it, in this case the calculations will not need to be changed.

We recommend internal validations where customers deviate from the stated protocol. As we have not performed this method using a 96 well plate for the final GOPOD assay therefore we cannot comment on its efficacy &/or resultant accuracy.