Aliquoting is the highest-return two minutes in peptide handling. It converts a stock that will degrade a little on every use into one that degrades once. The mechanics are simple; a few details determine whether it works as intended.
Size aliquots to the experiment, not to the tube
The point of an aliquot is that it is used completely and discarded. If your typical experiment consumes 40 µL, make 50 µL aliquots — not 200 µL, which guarantees three-quarters of each tube gets refrozen or thrown away.
It is worth spending a minute working out actual per-experiment consumption before pipetting. Aliquots that are slightly too small are a minor inconvenience; aliquots that are too large reintroduce exactly the problem being solved.
Choose tubes for adsorption, not for cost
Peptides adsorb to plastic, and cationic or hydrophobic sequences adsorb strongly. At high stock concentrations the loss is proportionally negligible. At working dilutions in the low microgram-per-millilitre range, adsorption can remove a meaningful fraction of the peptide before the experiment begins.
For dilute aliquots, low-binding polypropylene tubes are worth the cost. Glass is an alternative but carries its own surface chemistry and is less convenient at small volumes. Where adsorption is a known problem, a carrier protein such as 0.1% BSA in the diluent occupies binding sites — provided it does not interfere with the assay.
Fill volume and headspace
Leave modest headspace: aqueous solutions expand on freezing, and a completely full tube can pop its cap. Excessive headspace is also unhelpful, since the air-liquid interface is where interfacial denaturation happens. Roughly two-thirds full is a reasonable target.
Label so it survives the freezer
A label that becomes illegible at −80 °C is worse than no label, because it invites guessing. Cryogenic-rated labels or a solvent-resistant marker on the tube wall, not the cap — caps get swapped.
Record five things:
- Compound name
- Concentration
- Solvent or buffer
- Lot number of the source material
- Date of reconstitution
The lot number is the one most often omitted and the one that matters most a year later, when a result needs to be traced back to a specific certificate.
Freeze once, and quickly
Move aliquots to the freezer promptly after preparation. Rapid freezing produces smaller ice crystals and less time in the freeze-concentrated state. For valuable material, a brief plunge in dry ice or liquid nitrogen before transfer to −80 °C is a reasonable additional step.
Plan for losses
Two losses are systematic and worth building into the arithmetic.
Transfer loss. Pipetting a viscous or adsorptive solution repeatedly leaves material behind. Expect a few percent across an aliquoting session.
Dead volume. The last few microlitres of a tube are difficult to recover cleanly. Size aliquots with a small margin.
Set aside a single aliquot that is never used for routine work. If a batch of results ever comes into question, having untouched material from the original reconstitution — same lot, same day, one freeze cycle — is the difference between diagnosing the problem and speculating about it.