Protein estimation (Bradford method)

Standards preparation

10 mg/ml … 100 mg of BSA in 10 mL

• pipet 10 mL of sterile H₂O
• weight 100 mg (weight the weighing thing again - if something left)
• place BSA on to water surface; let slowly dissolve
• filtered 0.22 filter
• Store in -20°C

Dilution scheme for standarts:

[mg/mL]	mL to pipet [mL]
10	stock
5	0.5	from 10 mg/mL
2	0.2	from 10 mg/mL
1	0.1	from 10 mg/mL
0.5	0.5	from 1 mg/mL
0.25	0.25	from 1 mg/mL
0.1	0.1	from 1 mg/mL

• measure and prepare a standard curve
• express the coefficient of determination (R²)

General protocol

• pipette 1 µL of standard or sample; H₂O for blank
• add 999 µL of Bradford solution (diluted from concentrated stock 1:5 with H₂O)
• incubate 5 min; r.t.
• measure absorbance 595 nm

Compatibility
Ensure that the assay kit is compatible with your sample type, such as cell lysates, issue extracts, purified proteins, or peptides. Additionally, consider the compatibility with any additives or detergents present in your samples. Some common substances that potentially interfere with protein assay methods are reducing agents (e.g., DTT) and detergents (e.g., Triton X-100, SDS, NP-40).

For lysis buffers containing SDS of another detergent: BCA assay or Pierce

• pipette 1 µL of standard (prepared in H₂O or PBS) and add 1 µL of lysis buffer
• pipette 1 µL of sample (in lysis buffer) and add 1 µL of H₂O or PBS
• add 998 µL of reagent solution (diluted from concentrated stock 1:5 with H₂O)
• incubate 5 min; r.t.
• measure absorbance