Researcher organizing samples for a receptor assay

Wellness · Laboratory protocol

HCG

Human chorionic gonadotropin, a heterodimeric glycoprotein hormone (alpha and beta subunits with attached carbohydrate). Acts at the LH/CG receptor. Reference standard for endocrine and receptor-binding work.

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

LH/CG-receptor binding or cAMP accumulation in a cell line expressing the receptor, with a recombinant LH reference where subunit specificity is the question.

Required controls

Vehicle-only negative control
Receptor-null or antagonist condition
Validated reference material
Untreated viability control

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: This material is dosed in International Units, not milligrams. IU is a bioactivity unit, so IU-to-mass conversion is lot-specific and must come from the certificate, never from a general conversion factor.

Reconstitution concentrations

Solvent volume sets the stock concentration for this 5000 IU 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 5000 IU vial
Solvent added	Stock concentration	Per 0.1 mL	Per unit (U-100)
1 mL	5000 IU/mL	500 IU	50 IU
2 mL	2500 IU/mL	250 IU	25 IU
3 mL	1666.67 IU/mL	166.67 IU	16.67 IU
5 mL	1000 IU/mL	100 IU	10 IU
10 mL	500 IU/mL	50 IU	5 IU

Working range

Receptor-binding work is normally expressed in IU/mL or mIU/mL. Convert from the vial using the certificate potency for that lot.

LH/CG-receptor binding or cAMP accumulation in a cell line expressing the receptor, with a recombinant LH reference where subunit specificity is the question.

Stability

Lyophilized: Stable sealed and dry at the certificate temperature. Glycoproteins are more fragile than the small peptides here.
In solution: Noticeably less tolerant of freeze-thaw than the small peptides. Aliquot at first reconstitution.

Handling notes

Aliquot immediately at first reconstitution. Repeated freeze-thaw damages the heterodimer.
Do not vortex. Shear denatures glycoproteins in a way it does not affect a tripeptide.
Take the IU-to-volume calculation from this lot's certificate.
Avoid frothing during mixing; surface denaturation at the air-liquid interface is a real loss path here.

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.