Your peptide QC report consists of MS and HPLC analyses. The guide below will help you understand each analytical report, and how to tell if a quality analysis of your peptide was performed.
Even after multiple rounds of purification, small amounts of impurities may still exist in the final peptide product. Therefore, GenScript uses reversed-phase high performance liquid chromatography (RP-HPLC) to analyze the purity of each custom peptide.
RP-HPLC is a widely used analytical tool, in which the components of a complex mixture can be efficiently separated. The analyte mixture is usually dissolved in water, which is sometimes also mixed with an organic solvent or an acid to assist dissolution prior to RP-HPLC analysis. The analyte is carried by a mobile phase consisting of water and an organic modifier, which is pumped through a column packed with a stationary phase which usually is a number of small diameter particles consisting of carbon chains of a specified length on the surface. As the mixture is pumped through the column, the analytes (in this case your custom peptide and small impurities) adsorb to the hydrophobic surface of the stationary phase. As the percentage of organic modifier in the mobile phase is gradually increased, the analytes desorb into the mobile phase. Desorption of a specific analyte is based on its intrinsic properties, thus each analyte will remain in the column for a specific amount of time, called the retention time. Since peptide bonds maximally absorb UV light at the wavelength of 220 nm, a UV spectrometer is commonly used in RP-HPLC to detect a peptide as it elutes from the column. The detection signal is converted to a visual graph called a chromatogram, which is the plot of UV absorbance vs. elution time.
High resolution of eluted peptides is mainly based on the selection of columns and the organic modifier, especially its elution gradient. The height of a theoretical plate of each column used in the manufacture is regularly checked, and different gradients are established to separate varying peptides. Both processes are aimed at maintaining high resolution of every custom peptide. GenScript takes special care in developing RP-HPLC protocols to ensure the most effective separation and the accurate purity determination of your custom peptides.