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The BCA protein assay is a highly sensitive colorimetric method used to quantify protein concentration. It utilizes the color reaction of bicinchoninic acid with copper ions, which themselves have been reduced by peptide bonds in proteins, allowing quantification via absorbance measurement at 562 nm. Its resistance to common interfering substances makes it suitable for use with most biological samples.
Storage
Store at room temperature. Use within 1 year.
Note
For Research Use Only. Not for Diagnostic or Therapeutic Use.
Compatible substance concentrations in protein sample
Mix Reagent A and Reagent B at a ratio of 50:1 to prepare a working solution.
Dispense 25 µL of protein standards (e.g., a two-fold serially diluted BSA solution series starting at 2 mg/mL), blank (0 mg/mL), and samples into appropriate wells. Caution: Samples falling outside the calibration range must be measured again with appropriate dilutions.
Add 200 µL of the working solution to each well and mix gently.
Incubate at 37 °C for 30 minutes.
After incubation, remove any bubbles by gently tapping or shaking the plate and measure the absorbance at 562 nm (acceptable range: 540-570 nm).
Subtract the blank absorbance from all readings (blank correction).
Plot the blank-corrected absorbance values of the standard curve against protein concentration to obtain a regression equation.
Use this equation to calculate the protein concentration of each sample.
Protocol (Cuvette)
Mix Reagent A and Reagent B at a ratio of 50:1 to prepare a working solution.
Dispense 2.0 mL of the working solution into each of an appropriate number of sample tubes.
Add 100 µL of protein standards (e.g., a two-fold serially diluted BSA series starting at 2 mg/mL), blank (0 mg/mL), and samples to the respective tubes. Caution: Samples falling outside the calibration range must be measured again with appropriate dilutions.
Mix by vortexing or pipetting until thoroughly combined.
Incubate at 37 °C for 30 minutes.
After incubation, ensure that no bubbles are present in the light path of the cuvette, and measure the absorbance at 562 nm.
Subtract the blank absorbance from all readings (blank correction).
Plot the blank-corrected absorbance values of the standard curve against protein concentration to obtain a regression equation.
Use this equation to calculate the protein concentration of each sample.
H318 : Causes serious eye damage. H361 : Suspected of damaging fertility or the unborn child. H370 : Causes damage to organs. H372 : Causes damage to organs through prolonged or repeated exposure. H373 : May cause damage to organs through prolonged or repeated exposure. H317 : May cause an allergic skin reaction. H341 : Suspected of causing genetic defects. H410 : Very toxic to aquatic life with long lasting effects.
Precautionary Statements
P501 : Dispose of contents/ container to an approved waste disposal plant. P273 : Avoid release to the environment. P272 : Contaminated work clothing should not be allowed out of the workplace. P270 : Do not eat, drink or smoke when using this product. P202 : Do not handle until all safety precautions have been read and understood. P260 : Do not breathe mist or vapours. P201 : Obtain special instructions before use. P264 : Wash skin thoroughly after handling. P280 : Wear protective gloves/ protective clothing/ eye protection/ face protection. P302 + P352 : IF ON SKIN: Wash with plenty of water. P391 : Collect spillage. P308 + P311 : IF exposed or concerned: Call a POISON CENTER/doctor. P362 + P364 : Take off contaminated clothing and wash it before reuse. P333 + P313 : If skin irritation or rash occurs: Get medical advice/ attention. P305 + P351 + P338 + P310 : IF IN EYES: Rinse cautiously with water for several minutes. Remove contact lenses, if present and easy to do. Continue rinsing. Immediately call a POISON CENTER/doctor. P405 : Store locked up.
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