Procedure of standard Bradford assay:

1. Take out the frozen protein standard and allow it to come to room temperature.
   
   
2. As the concentration of the unknown protein sample can be anything, the assay will be performed with a range of dilutions (1, 1:10, 1:100, and 1:1000). Prepare 100 μl of each of the dilutions.
   
   
3. Take 15 test tubes and label them from 1 to 15.
   
   
4. Pipette out 10 μl , 20 μl , 30 μl , …….., 100 μl ovalbumin standard in the glass tubes labeled 1 – 10; leave blank the tube no. 11.
   
   
5. Add distilled water to make the final volume 100 μl in each of the tubes (including blank).
   
   
6. Take 100 μl of each of the unknown protein dilutions in the tubes labeled 12 – 15.
   
   
7. Add 5 ml of Bradford reagent in each of the tubes and mix well by inversion or gentle vortex mixing (avoid frothing).
   
   
8. Within 5 – 60 min , measure the absorbance of tubes 1 – 10 and 12 – 15 at 595 nm in the quartz/glass cuvette against the reagent blank (tube 11).
   
   
9. Record the readings in the suggested observation table below:

Observation Table:

Table 5.1: Observation table for the Bradford assay