MWGF200
Storage Temperature –20 °C
Gel filtration chromatography is an established method for determining the size and molecular mass of proteins. Fractionation is based on the diffusion of molecules into the pores of the resin. Larger proteins do not enter the pores of the resin as readily, but pass through the fluid volume of the column faster than smaller proteins. These protein molecules elute from the column in order of decreasing molecular mass.
The molecular mass determination of unknown proteins is made by comparing the ratio of Ve/Vo for the protein in question to the Ve/Vo of protein standards of known molecular mass (Ve is the elution volume and Vo is the void volume). The Vo of a given column is based on the volume of effluent required for the elution of a large molecule such as blue dextran (molecular mass of ~2,000 kDa, Catalog No. D4772). Plotting the logarithms of the known molecular masses of protein standards versus their respective Ve/Vo values produces a linear calibration curve.
Ve/Vo is essentially independent of column size and protein concentration, but may be temperature dependent for some proteins. Unreliable molecular masses may be obtained if the protein forms a complex with the gel, contains a large amount of carbohydrate, aggregates to larger complexes, or dissociates into subunits under the conditions used.1 The molecular mass of an impure protein may be determined using this procedure if a specific detection test is available for the protein.
The procedure for determining molecular masses using gel filtration chromatography as outlined in this bulletin is a modification of published methods.1,2 The protein standards in this kit may be suitable for use in other chromatographic systems including HPLC, although some buffer systems seem to alter the elution volumes of albumin (A8531) and carbonic anhydrase (C7025). The proteins in the MWGF200 Kit have a range of molecular masses from 12.4–200 kDa.
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This product is for R&D use only, not for drug, household, or other uses. Please consult the Safety Data Sheet for information regarding hazards and safe handling practices.
Store the kit at –20 °C.
Use of MWGF200 For Gel Filtration Chromatography
Figure 1.Typical calibration curve obtained with proteins from the MWGF200 Kit run on Sephacryl S-200-HR.
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