Protein Chromatography

Molecular Weight Markers for Gel Filtration Chromatography

Gel filtration chromatography is a well-accepted method for determining the size and molecular weight of proteins. The molecular weight of a given protein may be determined by comparing its elution volume with those of known protein standards. An essentially linear relationship is obtained if the logarithms of the molecular weights of standard proteins are plotted against their respective ratios of elution volume to column void volume, the column void volume being the elution volume of a very large molecule such as Blue Dextran.

Examples of typical proteins used for size determination include:

Protein Approximate Molecular Weight
Somatostatin 1640
Aprotinin, Bovine Lung 6,500
Cytochrome C, Horse Heart 12,400
Carbonic Anhydrase, Bovine Erythrocytes 29,000
Albumin, Bovine Serum 66,000
Alchol Dehydrogenase, Yeast 150,000
β-Amylase, Sweet Potato 200,000
Apoferritin, Horse Spleen 443,000
Thyroglobulin, Bovine 669,000
Blue Dextran 2,000,000

Sigma® supplies these proteins as individual proteins, as well as molecular weight range kits. Many of the protein standards are also suitable for use in other chromatographic systems, including HPLC. Some buffer systems seem to alter the elution volumes of Albumin and Carbonic Anhydrase.

Examples of calibration curves:

Calibration Curve Sephadex G-75-50 Graph Calibration curve typically obtained with specified proteins as run on Sephadex G-75-50. Calibration curve depicted should not be used to derive experimental results. Each laboratory must prepare its own calibration curves with its own system.


Calibration Curve Sepharose 6B Graph Calibration curve typically obtained with specified proteins as run on Sepharose 6B. Calibration curve depicted should not be used to derive experimental results. Each laboratory must prepare its own calibration curves with its own system.

References:

  • Whitaker, J.R. Anal.Chem., 35, 1950 (1963).
  • Andrews, P. Biochem. J., 91, 222 (1964).
  • Marshall, J.J. Chromatog., 53, 379 (1970).

Visit our listing of Molecular Weight Markers for Gel Filtration

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