Table 2. Comparison of different methods for determining protein concentration.
Instrument | Solution | Assay Method | Concentration |
Plate reader | BCA reagents | Abs at 562 nm | 103.9 ± 13.9 |
Spectrophotometer | H2O | Abs at 280 nm | 107.0 ± 6.9 |
Spectrophotometer | GnHCl | Abs at 280 nm | 94.2 ± 5.7 |
NanoDrop* | H2O | Abs at 280 nm | 88.0 ± 8.6 |
NanoDrop* | GnHCl | Abs at 280 nm | 98.1 ± 11.6 |
Concentration determination was performed using five dilutions (1∶1, 1∶2, 1∶5, 1∶10 & 1∶20) of a standard peptide solution (100 µM). The lowest dilution produced unreliable readings for all methods other than the BCA assay, and was not used for calculating the concentration or the standard deviation in those methods. Only the 1∶1 and 1∶2 dilutions produced reliable readings when using the NanoDrop; for this method, all other readings were omitted when calculating the concentration and standard deviation. * dilutions below 20 µM were unreliable and excluded.