COMPARISION OF DIFFERENT PROTEIN DETERMINATION METHODS
COMPARISION OF DIFFERENT PROTEIN DETERMINATION METHODS Company Method Detection Range Applications -Compatibility Assay protocol Precautions-Interferences A......
| Company | Method | Detection Range |
-Compatibility |
Assay protocol | |
| Absorbance 280nm | 0.1-1 OD280nm/ml | Absorbance of aromatic amino-acids (Trp, Tyr and Phe at less extent). Only for purified proteins with known absorptivity factor (Use ExPASy ProtParam tool to inquire E1% 280nm) | Read absorbance 280nm. | Nucleic acid, detergents, cofactors, phenolic compounds, pigments, reducing agents, etc etc. | |
| AMERSHAM- BIOSCIENCE (PHARMACIA) (# 80-6483-56) |
2-D Quant Kit | 0–50 µg
(1–50 µl) |
Designed for the accurate determination of protein concentration in samples prepared for electrophoresis and presence of detergents, Urea, DTT, EDTA, Ampholites, etc, and many buffer components. | Quantitative precipitation of proteins while leaving interfering substances behind. | |
| BIO-RAD (#500-0001/2/6) |
Bio-Rad Protein Assay (Modified Bradford) (pdf) |
0.2–0.9 mg/ml | Compatible with reducing agents (See list of compatible reagents on BioRad cataloge) |
Minimum incubation time 15minutes. Assay wavelength 650-750nm |
Detergents, basic buffers |
| BIO-RAD (#500-0111/2) |
DC Protein Assay (Modified Lowry) (pdf) |
0.1–2.0 mg/ml | Compatible with detergents, basic buffers (See list of compatible reagents on BioRad cataloge) | Minimum incubation time 15minutes. Assay wavelength 595nm |
Reducing agents |
| BIO-RAD (#500-0121/2) | RC DC Protein Assay (Modified Lowry) (pdf) | 0.1–2.0 mg/ml | Compatible with detergents, reducing agents, Laemmli sample buffer with 5% beta-mercaptoethanol, etc (See list of compatible reagents on BioRad cataloge) | Minimum incubation time 15minutes. Assay wavelength 650-750nm |
|
| GENO TECHNOLOGY (#786-005) | Non-Interfering Protein AssayTM | Compatible with reducing agents(2ME, DTT), chelating agents EDTA , detergents (non-ionic, anionic, cationic, and zwitterionic) , amines (Tris), sugar, urea, ammonium sulfate, guanidine hydrochloride, guanidine thiocyanate, drugs, antibiotics, cobalt, and numerous other agents. | Quantitative precipitation of proteins while leaving interfering substances behind. | ||
| MOLECULAR PROBES (N-6666) |
Nano Orange Protein Quantitation Kit (pdf) | 10 ng/mL -
10 µg/mL |
Little protein-to-protein variability. Compatible with the presence of reducing agents and nucleic acids. | Mix and heat 10' 95ºC. Fluorescence emissions are measured directly | Unusually high concentrations of lipids in the sample can interfere. This interference can be eliminated by acetone precipitation of the protein, followed by delipidation with diethyl ether. |
| MOLECULAR PROBES (C-6667) |
CBQCA Protein Quantitation Kit (pdf) |
10 ng/mL - 150 µg/mL | Functions well in the presence of lipids and detergents (to determine the protein content of lipoprotein samples or lipid–protein mixtures) | ||
| PIERCE (#23225) |
BCA Protein Assay (pdf) | 0.5-20 µg/ml | Compatible with detergents solubilized proteins. Proteins on affinity supports. Chaotropic agents, sugars, DNA, protease inhibitors | 2ml reagent + 0.1ml sample
Incubation 30' 37ºC. Read 562nm |
Reducing agents (can be eliminated with TCA, see Protocol) |
| PIERCE (#23235) |
Micro-BCA Protein Assay (pdf) | 0.5-20 µg/ml | Compatible with detergents solubilized proteins. Proteins on affinity supports. Chaotropic agents, sugars, DNA, protease inhibitors | 1ml reagent + 1ml sample
Incubation 60' 60ºC. Read 562nm |
Reducing agents (can be eliminated with TCA, see Protocol) |
| PIERCE (#23236) |
Coomassie Plus Protein Assay | 1-1500 µg/m | For quick estimation where accuracy is not important. Reducing agents. Chaotropic and chelating agents, metals, protease inhibitors. DNA. | 3 ml reagent + 0.1ml sample
Vortex and read 595nm |
Detergents (sometimes you can normalize using same detergent concentration in blank and standarts) |
| PIERCE (#23200)
SIGMA (#610-A) |
Coomassie Protein Assay (pdf) BRADFORD |
1-1500 µg/ml | For quick estimation where accuracy is not important: great variability. Compatible with reducing agents. Chaotropic and chelating agents, metals, protease inhibitors. DNA. | 5 ml reagent + 0.1ml sample
Vortex and read 595nm |
Detergents (sometimes you can normalize using same detergent concentration in blank and standarts) |
| PIERCE (#23240) |
Modified Lowry Protein Assay (pdf) | 1-1500 µg/ml | Accurate. Compatible with protease inhibitors. DNA. | 1 ml reagent + 0.2ml sample. Incubate 10' RT. Add 0.1ml Folin-Ciocalteu. Incubate 30' RT. Read 750nm | Reducing agents (can be eliminated with TCA, see Protocol,pdf) and some detergents |
| PIERCE (#23255) |
Fluoraldehyde Protein/Peptide Assay | 0.05-500 µg/ml | Compatible with reducing agents and some detergents. Chelating agents, metals and sugars | 2 ml reagent + 0.2ml sample. Mix and read fluorescence; excitation 330-390nm; emission 436-475nm. | Great variability. Cannot meassure in the presence of TRIS or Glycine buffer. |
| ROCHE (#1 767 283) (#1 767 003) |
ESL Protein Assay (pdf) | 20-800 µg/ml. Detection limit 1µg/sample |
Biuret-like reaction, detect peptide bonds. The assay is compatible with several detergents. Coul be used for determination of peptides and immobilized proteins | 7 minutes reaction. Read at 485nm. | |
| SIGMA (#690-1) |
BIURET | 1-10 mg/ml | Very accurate and simple | Read 550nm | Glucose, Ammonium sulphate, Sulphydryl compounds, PO4 buffers |
| Folin-Ciocalteu reagent: SIGMA (#F9252) - MERCK | LOWRY | 10 µg/ml - 1 mg/ml |
Very accurate | 1 ml reagent + 0.2ml sample. Incubate 10' RT. Add 0.1ml Folin-Ciocalteu. Incubate 30' RT. Read 750nm | Many detergents, reducing agents, EDTA, GuHCl, AmmSO4, >0.1M TRIS. Interfernce elimination with TCA or DOC-TCA |
| SIGMA (#P5656) Folin-Ciocalteu reagent: SIGMA (#F9252) - MERCK |
LOWRY-PETERSON | 1-10 µg protein | Modified Lowry for membrane proteins. Compatible with detergents and with the presence of all kind of interferents that can be eliminated by DOC-TCA precipitation. | DOC-TCA precipitation of proteins before Lowry assay | |
| BUTTERFLY (coomassie staining into 3MM Whatman paper) |
Very useful technique for proteins in all kind of solutions (like PAGE-SDS sample buffer) | Dry samples into 3MM Whatman paper.Treate with coomassie staining solution for ~30 min. Distain. Extracted ON with 3% SDS solution and read at 590 nm |
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发布日期：2008-2-3
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- Bradford protein assay
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