Researcher pipetting a copper-blue sample into a cell-culture plate

Cosmetic · Laboratory protocol

GHK-Cu

Copper(II)-binding tripeptide (Gly-His-Lys). Studied as a copper shuttle and in extracellular-matrix remodelling and fibroblast-signalling models.

For laboratory research use only. Material is supplied as a research reference standard and is not approved by Health Canada. Protocols below are compiled from published literature and supplier documentation; they are reference information, not medical advice. Anyone with a health condition or on medication should speak to a physician first.

Before you begin

• Lot-matched certificate and laboratory notebook
• Calibrated pipettes, low-binding tubes and suitable PPE
• Validated solvent, assay reagents and model-specific SOP
• Appropriate positive, negative and vehicle controls

Workflow

1Review the lot-matched COA and record identity, purity, batch number and received condition.
2Let the sealed vial equilibrate to room temperature before opening to limit condensation.
3Calculate the target stock concentration with C = mass / volume. Use only a solvent validated for the planned assay.
4Add solvent slowly down the vial wall, mix gently and avoid vortexing unless the validated method permits it.
5Prepare a log-spaced pilot concentration series; determine the usable range empirically instead of assuming a biological dose.
6Run the defined controls and technical replicates alongside every plate or experiment.
7Record preparation time, temperature, equipment, deviations and raw-data file locations.

Assay design

Fibroblast models with collagen synthesis, MMP/TIMP expression or migration endpoints. A copper-matched control (copper salt at equivalent concentration, no peptide) is required to separate peptide effects from free-copper effects.

Required controls

Vehicle-only negative control
Copper-matched control where relevant
Validated matrix-remodelling control
Parallel viability assay

Reported protocol

Injectable: 1.0 mg to 2.0 mg daily, in 30 to 60 day cycles
Topical: 1% to 2% solution applied daily to target skin or scalp areas
Route: Subcutaneous, or topical solution

Diluent

Sterile bacteriostatic water (0.9% benzyl alcohol) for multi-withdrawal stock, or sterile water for injection where a preservative-free stock is required by the assay.

Compound-specific: The reconstituted solution is deep blue. That colour is the copper complex and is expected, not a contamination signal. It does, however, interfere with any absorbance-based readout in the visible range.

Reconstitution concentrations

Solvent volume sets the stock concentration for this 50 mg vial. Lower volume gives a more concentrated stock and smaller working aliquots; higher volume gives a more dilute stock that is easier to measure accurately.

Stock concentration by solvent volume for a 50 mg vial
Solvent added	Stock concentration	Per 0.1 mL	Per unit (U-100)
2 mL	25 mg/mL	2.5 mg	250 mcg
3 mL	16.67 mg/mL	1.67 mg	166.67 mcg
5 mL	10 mg/mL	1 mg	100 mcg

Working range

Dermal-fibroblast work commonly spans the nanomolar to low-micromolar band.

Stability

Lyophilized: Stable sealed and dry at the certificate temperature.
In solution: Establish empirically. Protect from light.

Handling notes

Include a copper-only control at matched concentration. Without it, a copper effect reads as a peptide effect.
Blue colour interferes with colorimetric assays. Use a fluorescent readout, or subtract a peptide-only blank at the same concentration.
Avoid strong chelators in the buffer; they will strip the copper and change the molecule you are testing.

Documentation checklist

Record lot and COA, mass balance, solvent and concentration, preparation timestamp, storage history, plate map, replicate plan, instrument settings, exclusions and deviations. Preserve raw data and analysis code with the experiment record.