Assay¶

Definition¶

Assay is the quantitative determination of the amount or concentration of a peptide in a sample, typically expressed as a percentage of the labeled content or as weight/weight (w/w) basis.

Overview¶

While HPLC purity measures the relative proportion of peptide-related components, assay measures the actual amount of peptide present, accounting for water, residual solvents, counter-ions, and other non-peptide components.

Technical Explanation¶

Assay vs. Purity: - Purity: What fraction of the peptide-related material is the target peptide (area %) - Assay: How much target peptide is actually present in the total sample mass (w/w %)

Assay Methods: - UV spectrophotometry (using extinction coefficient) - HPLC with reference standard calibration - Amino acid analysis - Nitrogen determination (Kjeldahl/Dumas)

Factors Reducing Assay: - Water content (moisture) - Residual solvents - Counter-ion content (TFA, acetate) - Inorganic salts

Importance in Peptide Documentation¶

Assay data on the COA helps researchers accurately calculate the amount of active peptide for dosing. A product listed as "5 mg" may contain only 4 mg of actual peptide if the assay is 80% — the remaining 20% is water, salt, and counter-ion.

Frequently Asked Questions¶

Q: Why might assay be lower than HPLC purity? A: This is normal. HPLC purity reflects only peptide-related components. Assay is lower because it accounts for water, residual solvents, and counter-ions that are invisible to HPLC area normalization.

Q: What is a typical assay value for research peptides? A: Typical assay values range from 70% to 95%, depending on counter-ion type, water content, and salt form. TFA salts generally have lower assay values than acetate salts at the same purity.