To begin sample preparation for the Bradford protein assay, dilute the protein samples to obtain protein concentrations within the standard curve range of 0.025 to 1.0 milligrams per milliliter. Create multiple sample dilutions per sample within this range. For the zero protein point, add 10 microliters of the buffer or medium used to prepare the standard solutions to the designated wells.
Then add 10 microliters of each protein standard solution into a set of three wells serving as triplicates in a 96-well microplate. In a separate set of wells on the same 96-well microplate, add 10 microliters of each sample dilution in triplicates. Next, add 250 microliters of Bradford protein assay reagent to all wells.
A layout of the final microplate setup is displayed on the screen. Proceed to record the results in a well illuminated room within 5 to 15 minutes of adding the assay reagent. Using one hand, hold the microplate against a uniform white background, ensuring it is parallel to the work bench.
While using the other hand, hold a smartphone parallel to the bench and the microplate. Proceed to capture several pictures of the entire microplate. For iOS devices, enable the grid option in camera settings to turn on the camera level indicator.
For Android devices, activate grid lines in camera settings to achieve the same. Although no special lighting apparatus is required, maintain caution while capturing the images properly. Avoid shading the plate or the background and prevent excessive reflection.
Also, ensure not to tilt the plate. Inspect the picture for background, uniformity, shadows, and reflections. Also check the angle of the wells:The center of each well should be directly visible If desired, read the microplate in a microplate reader to compare the picture color data with the microplate absorbance readings.
