Thermo Scientific
Pierce BCA Protein Assay Kit
23227
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Quantitation of total protein in a sample is needed to be able to perform precise western blots. This kit was used to determine protein concentration at low concentrations.
Protein Concentration Quantitation
Total cell lysates of H4IIE rat hepatoma cells
There are two different procedures: one using test tubes and another using microplates. Our lab quantitates protein using microplates so we followed this procedure: Standards are made using diluent of sample and BSA that comes with the kit. The working reagent, based on bicinchoninic acid (BCA), is used for the colorimetric detection of protein and is diluted to a working range. 10uL standard and sample are used to obtain a working range of 125-2000ug/mL (25uL standard and sample can be used to obtain a working range of 20-2000ug/mL). 200uL of the working reagent is added to the microplate. The plate has to shake briefly and be incubated for 30 minutes at 37 degrees C. Absorbance is then measured on a plate reader in 540-590 range.
Note the 50:1 means 50 parts A and 1 part B.
The color visibly changes from light green to purple when protein is present. If reagents are pipetted correctly, the assay generates consistent standard curves which lower concentrations of protein can be quantitated reliably.
None
Works well with minimal variance
Use this product when protein concentrations are suspected to be minimal and assays with a higher working range are not compatible.