Protein Determination by the Warburg-Christian Method

1. OBJECTIVE

To standardize a procedure for the determination of protein content by Warburg-Christian.

2. SCOPE
This procedure applies to all products that require the determination of protein by Warburg-Christian method.

3. DEFINITIONS
Purified water – water from a deionizing system, resistivity > or = 18 MΩ•cm @ 25 °C

4. DISCUSSION
N/A

5. RESPONSIBILITIES
Analytical services laboratory personnel should follow this procedure as written.

6. SAFETY
Refer to Material Safety Data Sheets (MSDS) for hazards and appropriate handling precautions.

7. PROCEDURE

7.1 CONDITIONS:
T = 25 °C, A260 nm and A280 nm, Light Path = 1 cm

7.2 METHOD:
Spectrophotometric

7.3 REAGENTS:
Protein Sample Solution (Sample)

Prepare a solution containing approximately 0.3 mg protein/mL of sample in suitable buffer or purified water.

7.4 TEST METHOD
Measure the A_260nm and the A_280nm of sample in a suitable cuvette versus a suitable blank. The A_280nm must be greater than the A_260nm to obtain valid results. The presence of NAD or NADP will affect the A_260nm and A_280nm of proteins and thus, when necessary, must be corrected for in the calculations.

8. CALCULATIONS

mg protein/mL = [(1.31 x A280) - (0.57 x A260)] x dilution factor

*correction for NAD or NADP

9. REFERENCES & ATTACHMENTS
Warburg, O. and Christian W. (1941) Biochem. Z. 310, 384-421