Peptide reconstitution calculator: milligrams, millilitres, concentration per aliquot

A sealed vial of lyophilized peptide is a mass. The label says 10 mg, or 5 mg, or 80 mg for a blend. It says nothing about volume, because there is none: the cake at the bottom of the glass is dry.

Adding a solvent turns that mass into a solution, and a solution has a concentration:

C = m / V, where m is the mass of peptide stated on the label and V is the volume of diluent added.

Everything else on this page follows from those two letters. The mass is fixed by the supplier and confirmed by the lot certificate. The volume is the only variable the laboratory controls, and it is chosen so that the working aliquots come out at a volume the pipette handles well.

The calculator above also prints the mass carried by a 0.1 mL aliquot, because 0.1 mL is the smallest volume most bench pipettes measure without a visible error, and because reading the number in that form makes two vials of different strength directly comparable.

The whole table is one division, but it is quicker to read than to compute. Rows are the vial strengths sold as research reagents; columns are the volumes of diluent usually added. Each cell gives the concentration in mg/mL.

Vial	1 mL	2 mL	3 mL	5 mL
2 mg	2 mg/mL	1 mg/mL	0.67 mg/mL	0.4 mg/mL
5 mg	5 mg/mL	2.5 mg/mL	1.67 mg/mL	1 mg/mL
10 mg	10 mg/mL	5 mg/mL	3.33 mg/mL	2 mg/mL
20 mg	20 mg/mL	10 mg/mL	6.67 mg/mL	4 mg/mL
30 mg	30 mg/mL	15 mg/mL	10 mg/mL	6 mg/mL
50 mg	50 mg/mL	25 mg/mL	16.7 mg/mL	10 mg/mL
80 mg	80 mg/mL	40 mg/mL	26.7 mg/mL	16 mg/mL
100 mg	100 mg/mL	50 mg/mL	33.3 mg/mL	20 mg/mL

Multiply a cell by 0.1 to read the mass in a 0.1 mL aliquot. The 10 mg row brought up with 2 mL gives 5 mg/mL, so 0.5 mg per 0.1 mL. The 100 mg row (the size GHK‑Cu ships in) brought up with 5 mL gives 20 mg/mL, so 2 mg per 0.1 mL.

Three solvents turn up in a peptide laboratory, and they are not interchangeable.

• Bacteriostatic water: sterile water containing 0.9 % benzyl alcohol. The benzyl alcohol holds bacterial growth in check, which is what allows a vial to be opened more than once and kept for weeks in a refrigerator. This is the default for a peptide solution that will be sampled several times.
• Sterile water for irrigation or for laboratory use: no preservative. Once the septum is pierced the contents are a single-session preparation. Useful when the benzyl alcohol would interfere with an assay.
• Buffers and acidified water: some sequences dissolve poorly in plain water and go into solution in a dilute acetic acid or a phosphate buffer. The certificate or the supplier's data sheet says so.

The volume of the diluent vial is a separate number from the volume you add. OX ships 3 mL sealed vials: one vial covers one 3 mL preparation, or three preparations of 1 mL. The full description of the solvent, its benzyl alcohol content and its shelf life once pierced is in the bacteriostatic water guide.

Research use only. Not for human or veterinary use.

Two mistakes account for most of the discrepancy between the concentration a laboratory computes and the concentration it measures afterwards.

Fast addition. A jet of liquid aimed straight at the cake breaks it up and drives peptide onto the glass above the liquid line, where it stays. Running the diluent down the inside wall of the vial keeps the material in the solution. The cake then dissolves on its own in a minute or two; swirling helps, shaking does not.

Reading the wrong volume. The figure that matters is the volume of diluent added, not the volume the vial appears to hold afterwards. The freeze-dried cake occupies a small but real volume of its own, and a 3 mL vial that receives 2 mL reads slightly above 2 mL. The calculator uses the volume you added, which is the one you measured.

A vial that stays cloudy, or that leaves a visible deposit after several minutes, has not gone into solution. That is a property of the compound and the solvent, not of the arithmetic.

The calculator trusts one number: the mass printed on the label. That number is only worth the document behind it.

A lot certificate of analysis states the identity of the peptide, its purity by reverse-phase HPLC, its mass by mass spectrometry, and, on a full report, its net peptide content. The last field is the one that bears on this page. A vial filled with 10 mg of lyophilized material is not a vial holding 10 mg of peptide: the powder is an acetate salt and carries residual water, so the net peptide content is typically 80 to 90 % of the filled mass. A laboratory computing a precise concentration divides by the net figure, not by the label.

How to read each field of such a report is set out in how to read a peptide COA, and what a laboratory report proves (and does not prove) is in how to read a Janoshik test report. The Janoshik reports published per size are in the COA library.

A sealed lyophilized vial is stable for months. A solution is not, and the clock starts at the moment the diluent goes in.

• Sealed and dry: −20 °C for long-term storage, 2 to 8 °C for a few weeks, 24 months of shelf life from the date on the label.
• In solution with bacteriostatic water: 2 to 8 °C, protected from light, and the preparation is treated as short-lived. Benzyl alcohol limits bacterial growth; it does nothing against chemical degradation of the peptide itself.
• In solution without a preservative: a single session, or frozen in single-use aliquots so the same tube is never thawed twice.

Freeze-thaw cycles are the fastest way to lose a solution. Splitting it into aliquots at the moment it is made costs a minute and saves the batch. Compound by compound, the storage rules are in lyophilized peptides: storage and shelf life.