The certificate of analysis (COA) is the document that links a batch to the tests that characterise it. Knowing how to read it lets you decide whether a reagent is fit for an experiment before you use it.
1. Batch identification
The certificate states the batch number, the analysis date and the laboratory that ran the tests. The batch number must match the label on the vial you received.
2. Identity: mass spectrometry
Electrospray ionisation mass spectrometry (ESI-MS) measures the mass-to-charge ratio. Peptides usually appear as multiply charged ions, for example [M+2H]²⁺ and [M+3H]³⁺, and the software reconstructs the molecular mass. Compare the observed mass with the theoretical one:
- Differences of a few tenths of a dalton are normal.
- +16 Da usually indicates oxidation, typically of methionine.
- −18 Da indicates water loss.
- A difference equal to the mass of one residue suggests a deletion.
3. Purity: HPLC
Reversed-phase liquid chromatography separates the peptide from its impurities. Detection at 214 nm records the peptide bond, so it responds to almost every peptide species. Purity is reported as main-peak area over total area: 99 % means that 99 % of the 214 nm signal belongs to the target peptide. It is not the percentage of peptide in the powder.
- The chromatogram should show one sharp main peak and its retention time.
- Small peaks before the main one are usually truncated, more hydrophilic sequences.
- Peaks right next to the main one may be D-isomers or deletions that are hard to separate.
4. Net peptide content
Besides the peptide, the lyophilisate contains counter-ions (TFA or acetate) and water. If the certificate reports net peptide content, use it to calculate working concentrations.
5. Endotoxins
They are measured with the LAL assay and expressed in endotoxin units per milligram (EU/mg). The lower the value, the better for assays with immune cells. Check the limit your protocol requires.
6. Document integrity
When the original laboratory certificate is published on wwup., its SHA-256 hash appears next to the batch. If you compute the hash of the downloaded file and it matches, the document has not been modified:
sha256sum COA-WW-BPC-402-laboratorio.pdf # Linux or macOS
certutil -hashfile COA-WW-BPC-402-laboratorio.pdf SHA256 # Windows