Bradford assay for protein quantification (TKs purified)

A stock solution of BSA (Bovine serum Albumine) 1 mg/ml is made and from this solution different aliquots are picked in order to obtain different dilution solution.

1. 25 µL from solution + 975 µL water to obtain 25 µg/ml
2. 50 µL from solution + 950 µL of water to obtain 50 µg/ml
3. 75 µL from solution + 925 µL water to obtain 75 µg/ml
4. 100 µL from solution + 900 µL water to obtain 100 µg/ml
5. 125 µL from solution + 825 µL water to obtain 125 µg/ml
6. 150 µL from solution + 850 µL water to obtain 150 µg/ml
7. 200 µL from solution + 800 µL water to obtain 200 µg/ml
8. 250 µL from solution + 750 µL water to obtain 250 µg/ml
9. Stock solution 1 mg/ml The solutions are assessed in triplicate A 96 well microplate assay is prepared where each well is filled in with 160 µL of Bradford solution G250. Then 40 µL of the 8 diluted solutions are added to 8 different wells in order to obtain a calibration curve. The solutions are gently mixed in order to avoid precipitation and bubble wraps and then the plate is left in dark at room temperature for 15 minutes. Meanwhile a protein’s sample solution of 4 mg/ml is prepared and diluted (5 times, 10 times and 20 times) and 40 µL are added to a new well. The same procedure is followed for all the proteins 4 mg in 1 ml then 50µL are picked from this solution and diluted with 200 µL water then 50 µL are picked from the stock and diluted with 450 µL water then 50 µL are picked from the stock and diluted with 950 µL water