Reconstitution math is one of the most common sources of error in laboratory peptide work. This guide walks through the calculations clearly, using practical examples and a reusable formula.
The Core Formula
Every reconstitution problem reduces to a single relationship:
Rearranged for the two most common scenarios:
- To find concentration: Concentration = Mass ÷ Volume
- To find required volume: Volume = Mass ÷ Desired Concentration
- To find mass per aliquot volume: Mass per aliquot = Concentration × Aliquot Volume
Worked Example 1: Standard Reconstitution
You have a vial containing 5 mg of peptide. You want a working concentration of 5 mg/mL.
- Volume of solvent to add = 5 mg ÷ 5 mg/mL = 1.0 mL
Add exactly 1.0 mL of bacteriostatic water (or other appropriate solvent) to the vial. Final concentration is 5 mg/mL throughout the vial.
Worked Example 2: Calculating What’s in a Given Volume
Same vial: 5 mg total, reconstituted with 1.0 mL of solvent. The final concentration is 5 mg/mL, which equals 5,000 micrograms per milliliter (5,000 mcg/mL).
| Volume Withdrawn | Mass of Peptide |
|---|---|
| 1.00 mL | 5,000 mcg (5 mg) |
| 0.50 mL | 2,500 mcg |
| 0.25 mL | 1,250 mcg |
| 0.10 mL | 500 mcg |
| 0.05 mL | 250 mcg |
Worked Example 3: Aliquot Mass from Volume
For laboratory aliquoting and dilution math, the relationship between withdrawn volume and peptide mass is linear. If your reconstituted concentration is 5 mg/mL:
- 0.10 mL aliquot = 500 mcg of peptide
- 0.20 mL aliquot = 1,000 mcg (1 mg) of peptide
- 0.50 mL aliquot = 2,500 mcg (2.5 mg) of peptide
The general rule: mcg per aliquot = concentration (mcg/mL) × aliquot volume (mL). For a 5 mg (5,000 mcg) vial reconstituted in 1.0 mL of solvent, each 0.01 mL aliquot contains 50 mcg.
Worked Example 4: Different Vial Size, Different Concentration
You have a 10 mg vial and want a final concentration of 2 mg/mL.
- Volume to add = 10 mg ÷ 2 mg/mL = 5.0 mL
If your vial only holds 3 mL, you have a problem — your desired concentration is too dilute for the container size. Either choose a higher concentration (3.33 mg/mL with 3 mL of solvent) or split into multiple vials.
Quick Reference Table
For a 5 mg peptide vial, here is how the math changes with different solvent volumes:
| Solvent Added | Final Concentration | Mcg per 0.10 mL aliquot |
|---|---|---|
| 1 mL | 5 mg/mL | 500 mcg |
| 2 mL | 2.5 mg/mL | 250 mcg |
| 2.5 mL | 2 mg/mL | 200 mcg |
| 5 mL | 1 mg/mL | 100 mcg |
Best Practices
- Pick a clean concentration. Whole-number concentrations (1, 2, 5, 10 mg/mL) make downstream math far easier and reduce calculation errors.
- Write the math on the vial. Label each vial with date, mg in vial, solvent volume, concentration, and any conversion (e.g., “5 mg/mL = 500 mcg per 0.10 mL”).
- Use a calibrated volumetric instrument. Inexpensive laboratory syringes and pipettes have volume markings that vary by ±5–10%. Calibration matters for reproducible bench work.
- Double-check before drawing. A 10× error in concentration math is the single most common reconstitution mistake.
Solvent Choice Quick Reference
- Bacteriostatic water (0.9% benzyl alcohol): General-purpose laboratory solvent. Supports multi-aliquot storage of reconstituted solutions in refrigeration for laboratory use.
- Sterile water (laboratory grade, preservative-free): Used as a solvent when bacteriostatic preservatives are incompatible with the specific sequence or downstream assay. Short shelf life once opened.
- 0.1% acetic acid or dilute acid: Used for some basic peptides that resist dissolution in neutral water.
- Dilute ammonium hydroxide or basic buffer: Used for some acidic peptides.
Always consult the sequence-specific solubility data on the Certificate of Analysis or solubility profile sheet before choosing a non-standard solvent.