Pure albumin and its method of preparation and detection
The subject of the present invention is a pure monomeric bovine serum albumin, a method of producing it characterised by the use column chromatography in resin and a method of identifying it using dynamic light scattering.
1. A method of obtaining analytically pure monomeric bovine serum albumin devoid of high molecule weight protein aggregates and saccharide contaminates comprising:
a) obtaining a bovine serum albumin preparation, and
b) purifying the preparation obtained in step a) by performing chromatography on a HW-55 resin equilibration with a buffer, so as to obtain analytically puer monomeric bovine serum albumin devoid of high molecular weight protein aggregates and saccharide contaminants.
2. The method according to claim 1 , wherein step a) includes removing hydrophobic ligands from the bovine serum albumin.
3. The method according to claim 1 , wherein the chromatograpy of step b) is performed at room temperature at a rate of 1.2 ml/10 min.
4. The method of claim 1 , further comprising step c) after step b), wherein step c) comprises:
c) performing dynamic light scattering to determine the diameter of bovine serum albumin molecules in the sample obtained from the chromatography of step b), wherein the presence of bovine serum albumin molecules with a diameter of about 7 nm indicate the sample contains analytically pure monomeric bovine serum albumin devoid of high molecular weight protein aggregates and saccharide contaminants.
5. The method of claim 1 , wherein the buffer of step b) is 0.1 M acetate buffer, at pH 5.65, containing 1% n-butanol.
6. The method of claim 2 , wherein the hydrophobic ligands are removed by passing the bovine serum albumin through charcoal.
7. The method of claim 1 , wherein the monomeric bovine serum albumin is analytically pure and devoid of high molecular weight protein aggregates and saccharide contaminants as determined electrophoretically by SDS/PAGE in a 12% separating gel.
8. The method of claim 7 , wherein the monomeric bovine serum albumin is devoid of saccharide contaminants as determined by phenol method analysis.