In this informative blog, we will go over the concentration of low molecular weight!
It is typically helpful and desirable to reduce the amount of a protein sample to conduct further analysis. Some applications require the protein or peptide to be fully contained in a small volume. For example, plenty of protein purification experiments need the eluate from a specific method to be concentrated to help enable the sample to be tested for activity in some tissue culture assay. Similarly, column eluates to be run on PAGE gels has to be concentrated to a tiny volume for loading. Other applications require the recovery and concentration of peptides, such as the products of protein proteolysis experiments where the peptides are needed for sizeable spectroscopic microsequence analysis. In this informative blog, we will go over the concentration of low molecular weight!
Methods
Four milligrams of Insulin A chain, or Molecular weight 2.5 kDa, and 4 mg of thyrocalcitionin, or Molecular weight 3,593 Da, were made up in 4ml of 20 mM phosphate buffer. Peptide solution was added to 3x Vivaspin 500 ultrafiltration devices incorporating a three kDa MWCO PES Molecular weighing cut-off membrane. The samples are then centrifuged at 13,000 rpm for one hour at room temperature or at least until the retentate is equal in volume in 100.
Protein Estimation
The protein content of the filtrate and the retentate for both Insulin A chain peptides and thryocalcitonin were measured using an Advanced Protein assay.
The Results
The protein content of the filtrate and retentate for both Insulin A chain peptide and thyrocalcitonin following the concentration of both proteins within the Vivaspin 500 are shown in Table 1. The percentage of recovered protein in the retentate is offered as well.
The Main Conclusions
It is clear from the results presented that the Vivaspin 500 is a reliable and efficient method to use in the lab for the concentration of peptides. These devices are handy for the concentration of and the exchange of buffer. Any sample that might require such manipulations is precious since the recovery from the main rentate of the Vivaspin 500 is incredible.
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