Freezing Starter-Sized Samples of Yeast for Long-Term Storage
Written by Brewitt with contributions from HBT members
WHAT: The purpose of this article is to summarize what has been learned about freezing yeast by a number of contributors to the thread "Do you know how to make a yeast starter? Then why not farm yeast and freeze it?" (http://www.homebrewtalk.com/f163/do-you-know-how-make-yeast-starter-then-why-not-farm-yeast-freeze-269488/) started by, and with extensive contributions from, BBL_Brewer. Other major contributors include Brewitt (me), Forkhead, katy bug, ScoRas, and IsItBeerYet. Many others have made contributions that can be read in their original form in the thread. Together, through reading and experimentation, we have come up with an effective and relatively simple method for long term storage of pitchable quantities of yeast in either standard frost-free freezers or, better yet, non-frost-free freezers often used for food storage. Keep in mind that some of the recommended procedures have not been fully tested and will undoubtedly be undergoing some modifications but the method outlined below works well. Some modifications and additional considerations are listed at the end of the article.
WHY: The method(s) described below is sufficient for storing cells in portions large enough to pitch directly (50-200 billion cells, similar to the amount in a smack pack from Wyeast or a tube from White Labs) (See Figure 1). Nevertheless, we recommend an overnight starter to ensure a large number of viable cells and rapid initiation of fermentation. The cells to be frozen can be from a large starter culture made from a tube or pack of commercial yeast. A single large starter can be broken up and saved for multiple future brewing days. A similar approach can be taken with a small starter made from yeast recovered from commercial bottle-conditioned beer or home brews that you would like to save for later. Finally, the yeast can be washed cells from a recently completed fermentation (a single 5 gallon fermentation can provide starter samples for as many as 10 future brews). Keep in mind that you will save $7-$10 per 5 gallon batch of beer each time you pull a tube out of the freezer.
GOALS: The goals of the method that we have devised are three-fold:
- To allow the preservation of large samples of cells in freezers in common use by homebrewers
- To use materials available to homebrewers at a reasonable cost
- To achieve high viability of stored cells for weeks, months, or perhaps years
SUPPLIES: Materials required for preparation of frozen cells:
- A large quantity of yeast culture that you want to store for future use. Any "clean" yeast culture can be used (for example: one or more quarts/liters of aerated or stirred starter culture; washed cells from a 5 gallon fermentation (see "Sticky: Yeast Washing Illustrated") (http://www.homebrewtalk.com/f163/yeast-washing-illustrated-41768/)).
- Sanitized tubes for freezing samples of yeast. This could be plastic White Labs tubes saved from prior purchases (these are apparently also called "baby soda bottles"); 50 ml plastic screw top tubes commonly used in laboratories (see Resource section at the end of this article), any other small freezable container that can be sanitized and tightly sealed. See Figure 2.
- Some form of tube holder.
- Glycerin, food grade (must be diluted in sanitized water before use; a rate of 1 part glycerin to 4 parts water gives a 20% final concentration). Glycerin, glycerine and glycerol are all the same thing. See Resource section at the end of this article. See Figure 3.
- Isopropyl alcohol (also called "rubbing alcohol", available at any drug store), freezable gel packs, other good insulator for tubes of yeast.
- Container large enough to hold multiple tubes of yeast that will hold alcohol or gel packs. An insulated container is best (small Styrofoam ice chest or discarded shipping box for frozen goods).
- Sanitized water
HOW: A single method is outlined. Alternatives are discussed in below.
1. Collect yeast by chilling the culture or washed cells in a refrigerator without disturbing for 4 hours or more (preferably overnight). Pour off spent wort or washing water leaving just enough to allow cells to be suspended as a very dense slurry. The goal is to get sufficient cells for an overnight starter culture in a volume that is convenient to freeze. A good sample size is about 50-200 billion yeast cells (0.5-2 x 1011 cells). One quart of starter culture can usually be reduced to 1/40th the volume or less giving you about 100 billion cells in 25 milliliters (about 1 1/2 Tablespoons).
2. Split up the slurry of settled yeast into samples of about 100 billion cells (cells from about 1 quart of starter or a half gallon of fermented beer) in sanitized freezable screw top containers. That amount of slurry should fill the container less than half way.
3. Add an equal volume of chilled 20% glycerol to each of the samples of yeast and cap tube tightly. Swirl or stir until fully mixed without frothing. This will lead to a final concentration of glycerin of 10%. Concentrations of glycerin higher than 15% appear to be detrimental to cells when stored at home freezer temperature (20C, -3F). Make sure there is sufficient room for expansion during freezing.
4. Loosen the caps of the tubes and place into the large container, standing up to avoid spilling, and add enough isopropyl alcohol to the large container submerge the tubes to the level of the glycerin solution but not to the level of the cap. You don't want isopropyl alcohol mixing with your yeast. If using gel packs, make sure tubes are completely surrounded by the gel.
5. Place the container in the freezer. The isopropyl alcohol or the gel packs will slow the freezing of the cells, which is an advantage for maintenance of viability, and will avoid thawing during frost-free cycles.
6. After freezing is complete, tighten the caps of the tubes. If you are in a non-frost-free freezer, you can remove them from the alcohol and save it for a future use.
1. Remove tube of yeast from the large freezing/storage container.
2. Immediately submerge tube in water at approximately body temperature (37C, 98F). This can be running warm water or a large volume of water for submersion of the tube. Swirl continuously until completely thawed.
3. Remove tube of cells from the water, wipe dry with sanitizer and pitch into 1-4 liters of wort to prepare an overnight starter. In general that inoculation of the starter will grow to maximum density within 12-24 hours with appropriate shaking or aeration.
Other useful considerations and information:
Keep in mind that the cleanliness of your starter culture, washed yeast, and freezing supplies is key. Your subsequent beers will only be as good as your starting yeast strain and any contamination in your starter culture will be perpetuated in brews made with the frozen yeast prepared from it. If you have a beer with flaws due to contamination, don't freeze the yeast for later use. Also, yeast blends and blends of yeast and bugs will not necessarily be present at the same ratios after growing starters or using them for a fermentation. They can be frozen but results may vary.
Isopropyl alcohol for freezing and for storage can be cumbersome. The goal is to freeze slowly and avoid thawing during frost-free cycling in home freezers. Another approach to slow freezing is to chill in a refrigerator overnight and then to place in an insulated container like a Styrofoam ice chest or box and place in the freezer. It is useful to use gel packs or some similar frozen insulator when storing in a frost-free refrigerator.
Glycerin is a cryoprotectant. It helps maintain the viability of frozen cells. We have tested final concentrations from 7.5% to 50% and found 7.5%-15% to be optimal for maintaining viability (upwards of 75% viability after several months of freezing). Lower final concentrations may also work but higher concentrations are detrimental.
Glycerin can be added to the yeast slurry before or after splitting into smaller portions. If you want a more concentrated slurry, you can pour off more spent wort and/or resuspend your slurry in more concentrated glycerin to achieve a final concentration of 7.5%-15%.
If you have yeast that have been settled at room temperature, you can add glycerin solution that is at room temperature and then chill.
Although yeast can be used directly, this is likely to lead to under-pitching and a slow start to fermentation.
Resources: (The following are meant as examples, not necessarily recommended vendors or products)
Tubes for freezing yeast: Some examples from Amazon
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