Bradford Assay

Pipet 98 microL of 1X PBS into a labeled microcentrifuge tube, then add 2 microL of milk sample. Mix thoroughly by pipetting or vortexing to create a 1:50 dilution.

Label two cuvettes as 'control' and 'sample'. Pipet 20 microL of diluted milk sample into the sample cuvette and 20 microL of 1X PBS into the control cuvette, being careful not to touch the optical surfaces.

Label cuvettes for each protein standard with corresponding concentrations marked clearly. Add 20 microL of each protein standard into the appropriate cuvette using clean pipet tips for each sample.

Add 1 mL of 1X Bradford reagent to all cuvettes (sample, control, and standards) and mix completely by pipetting up and down. Incubate at room temperature for 5 minutes.

After 5 minutes, compare the color of the milk sample cuvette against the protein standard cuvettes and determine which standard most closely matches. Estimate the protein concentration based on this visual comparison.

Insert the control cuvette into the spectrophotometer and use it as a blank to set the instrument to zero absorbance or 100% transmittance.

Read and record the absorbance values for all seven protein standards in the spectrophotometer according to protocol instructions.

Remove the blank cuvette and insert the milk sample cuvette into the spectrophotometer. Read and record the absorbance value.