Same lesson, with the figures in motion.
Preserve Strains
Why freeze a strain?
This is not a lesson you need for every brew. But it is worth an evening the day you have a strain you would hate to lose — a wild yeast you caught well, a strain that has become hard to find, or simply your favourite.
1. The Cryoprotectant
Water that freezes unprotected forms sharp, crystalline needles of ice. They grow straight through the cell membrane and burst the cell from the inside — the yeast does not die of the cold, it dies of the ice.
Glycerol solves this by working its way into the cell and lowering the freezing point. The liquid then sets not into a hard block of ice crystals but into a viscous, glassy mass with no sharp edges. The cell's inner structures come through the whole winter in the freezer unharmed.
The protocol uses a 40% glycerol solution mixed 1:1 with thick slurry, which gives about 20% in the finished vial — the concentration that gives the best cell survival without becoming toxic to the yeast.
2. Boil the Solution
Mix 40 mL glycerol (50 g) + 60 mL water in a glass bottle — that gives a 40% glycerol solution. Screw the cap on loosely.
Glycerol is viscous and clings to the measuring cylinder. Weigh it instead of measuring by volume: 1 mL of glycerol weighs about 1.26 g.
Pressure cook for 15 minutes at 121 °C and let it cool to room temperature with the cap still on. No pressure cooker? Boil the bottle in a water bath for 20 minutes with the cap loose — weaker protection, so use the pressure cooker if you have one.
The solution keeps for months in the fridge, so make the whole batch at once.
3. Filling the Cryovial
Sterilise the vials in advance (pressure cook them empty with the caps loose) or use ready-sterile cryovials. Unscrew the cap and stand the vial in a rack — never hold it in your hand while you dose.
Add 0.5 mL of thick yeast slurry and 0.5 mL of the 40% solution — equal parts, which give you those 20 percent. One millilitre in total in a 2 mL vial: plenty of margin to the protocol's limit of two thirds at most, because the liquid expands as it freezes.
Turn the vial upside down a few times and let it stand for 15–30 minutes at room temperature. The glycerol needs that time to work its way into the cells — skip the rest and you are freezing unprotected cells.
Label the vial with strain, date and generation using a freezer-proof pen. One vial gives you one wake-up later, so count on 8–10 vials per strain for many years of supply.
4. The Freezer
Stand the vials in the freezer at −18 to −20 °C and do not disturb them again. Right at the back and right at the bottom the temperature is steadiest — not in the door.
Their defrost cycle lets the temperature swing up and down, and every swing builds fresh ice crystals in the vial. A manual freezer without defrost is far better — if all you have is a frost-free one, stand the vials in an insulated box (cool bag, polystyrene) inside the freezer to soften the swings.
Count on several years of shelf life at −18/−20 °C. A lab's −80 °C is another league entirely (decades), but an ordinary household freezer is plenty for a home library.
Label the box clearly — the glycerol vials look like food, but they are not.
5. Waking a Strain
Take out one single vial and close the freezer straight away — the others must not start thawing at the edges.
Thaw it quickly in your hand or in lukewarm water, just until the contents turn liquid. Do not let it stand and get warm.
Pour the whole contents into a small sterile starter culture — around 100 mL of weak wort (1.020–1.030) at room temperature. No streaking onto a plate: the yeast you caught goes straight into liquid where you can see that it is alive.
Be patient. A frozen culture can need 24–72 hours before anything shows — do not write the vial off too early. Once it is going you step it up like any other starter.
Freeze-thaw cycles kill yeast faster than anything else in this process. If you have plenty in stock: freeze fresh vials next time you have a healthy culture running anyway, with the next generation number on the label.