Net Peptide vs. Gross Peptide Weight
When ordering or receiving quotations for peptides, it is important to specify the amount in either gross or net peptide weight. In most cases, peptides are sold in gross peptide weight unless otherwise requested.
After purification and final lyophilization, the white peptide powder will contain some counter ions, residual trace solvents, and moisture. The most common counter ions include trifluoroacetate, sodium (Na+), and acetate (OAc-). If a TFA buffer was used during the final purification, for example, it can form a salt with amino groups on the side chains of lysine and arginine residues or on the amino terminus of the peptide. TFA salt, the residual traces of TFA, can cause the pH of your solution to drop below 7. For this reason, it is often necessary to check and adjust the pH of your peptide solution to reach your desired pH. In addition to counter ions, peptides can absorb and maintain a particular amount of moisture or water content. This depends primarily on the hygroscopic nature of the molecule. As a general rule, the greater the percentage of hydrophilic residues contained in your sequence, the more hygroscopic the peptide will be. It is generally impossible to render a peptide completely anhydrous.
When calculating the concentration of peptide solution for your biological assays it is essential that you account for peptide content. The peptide content can be determined by amino acid or nitrogen analysis for each lot of material. It is important to note that the peptide content can vary from lot to lot. To insure consistent peptide concentrations, subtract the non-peptide weight when determining what volume of solvent to dissolve your peptide. For example, to make a 1mg/ml solution of peptide with a content of 80%, you would use 800ul of solvent instead of 1000 ul.