everty2007: If you wash yeast, there are only a few more steps to the process. Settle the yeast and pour off the chilled water. Then I would add cold glycerine diluted to give the appropriate final concentration when combined with the yeast slurry. Finally, freeze. Use some form of protection from thawing if you are putting them in a frost free freezer. I don't think there is much to video.
Freezing Yeast
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Jersh: Sounds like we have the same training and resources. Let me clarify. The goal here is not to prevent freezing. 50% glycerol will not freeze at home freezer temperatures. 10-20% will. That is the range that we are shooting for. I used 20% but others say between 10-15% is optimal for recovery of cell viability using this approach. I freeze mine in an ultracold (-80C) which, of course, has even better preservation properties. Use 10-20%. Even in an ultra cold, where 50% will freeze, 20% yields better viability. I hope that answers your question.
With regard to settling of the yeast: Yes, if you freeze in a home freezer the yeast is going to settle some before it freezes. This is not a problem. The point of the glycerol is not to suspend the yeast, it is to affect the osmolarity of the yeast and the medium. That happens before the yeast freeze. So, no problem.
With regard to settling of the yeast: Yes, if you freeze in a home freezer the yeast is going to settle some before it freezes. This is not a problem. The point of the glycerol is not to suspend the yeast, it is to affect the osmolarity of the yeast and the medium. That happens before the yeast freeze. So, no problem.
everty2007
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@Brewitt I picked up all the supplies today and will be attempting this tomorrow.
Zoidberg
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@Brewitt
I am going to be trying this tomorrow. I have a few questions.
When sanitizing water should I add glycerol to the boil or add after? Will boiling effect the glycerol?
I am using a home freezer and don't have room for a styrofoam cooler. I am using the 50ml Centrifuges like you have. I set up a jar with a tin foil lid and pushed the centrifuge through and plan to fill jar with rubbing alcohol. Will this be sufficient to keep it from thawing/re-freezing or would buying some gel packs and tightly wrapping be a better idea?
I am going to be trying this tomorrow. I have a few questions.
When sanitizing water should I add glycerol to the boil or add after? Will boiling effect the glycerol?
I am using a home freezer and don't have room for a styrofoam cooler. I am using the 50ml Centrifuges like you have. I set up a jar with a tin foil lid and pushed the centrifuge through and plan to fill jar with rubbing alcohol. Will this be sufficient to keep it from thawing/re-freezing or would buying some gel packs and tightly wrapping be a better idea?
@Brewitt, Thanks for the tutorial.
You want to sanitize the glycerol so you can put dilute it to working concentration in a bottle and put the bottle in boiling water for 15 minutes. A canning Jar is good for that.
The bottle sounds like a great solution for storing the tubes. Or just a wide mouth bottle you can throw a couple tubes in and put something that doesn't freeze like alcohol or gel pack. Close the tubes tightly.
You might get more ideas from the thread on the forums.
The bottle sounds like a great solution for storing the tubes. Or just a wide mouth bottle you can throw a couple tubes in and put something that doesn't freeze like alcohol or gel pack. Close the tubes tightly.
You might get more ideas from the thread on the forums.
![Craft A Brew - Safale S-04 Dry Yeast - Fermentis - English Ale Dry Yeast - For English and American Ales and Hard Apple Ciders - Ingredients for Home Brewing - Beer Making Supplies - [1 Pack]](https://m.media-amazon.com/images/I/41fVGNh6JfL._SL500_.jpg)
You want to sanitize the glycerol so you can put dilute it to working concentration in a bottle and put the bottle in boiling water for 15 minutes. A canning Jar is good for that.
The bottle sounds like a great solution for storing the tubes. Or just a wide mouth bottle you can throw a couple tubes in and put something that doesn't freeze like alcohol or gel pack. Close the tubes tightly.
You might get more ideas from the thread on the forums.
The bottle sounds like a great solution for storing the tubes. Or just a wide mouth bottle you can throw a couple tubes in and put something that doesn't freeze like alcohol or gel pack. Close the tubes tightly.
You might get more ideas from the thread on the forums.
Zoidberg
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Ok thanks so I think what I will do will be incredibly rigged but totally work. The bottom half of a 2 litter bottle with cardboard cut to fit the top, and about 6-7 hoes cut to fit 50ml Centrifuge tubes. I will fill with alcohol, put cardboard over and tape loosely on edge with electric tape. Then drop tubes of yeast in through hols and lay 2 gel packs on top of them. This will all be set inside of my large ice tray filled with ice.
enohcs
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As a VWR rep...I APPROVE THIS ARTICLE AND ALL THE PHOTOS OF MY PRODUCT
user 47131
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Maybe I am missing something very simple here, but what is the actual quantity of glycerine being added to the vials? Maybe I'm confused by the fact that we are already watering the glycerine down to a 20% concentration (also, based on the 1/4 ratio, aren't we really starting out with a 25% concentration?), so to achieve that optimal 15% concentration overall, how do we figure that? Thanks.
stpug
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@keesimps
It's all in the math
I have 50ml vials.
I create a 50% glycerin solution because it's easy to mix.
I want 15% glycerin concentration in my vials.
15% of 50ml is 7.5 ml of GLYCERIN.
Double that to 15ml because my stock solution is only 50% glycerin.
(50ml * 0.15 / 0.50 = 15 ml of 50% glycerin solution to 35 ml yeast slurry)
I can then fill the vial up with any amount of anything (except glycerin) and maintain a 15% glycerin concentration in my vial. Regardless of how thick/thin my yeast slurry is, as long as I fill up to 50ml the 15% glycerin concentration is maintained. If I'm a little short in one vial then I top up with sterile water to the 50ml mark - no problem.
The hard/questionable part is estimating how many billion cells you have put into each vial AND it's viability once thaw.
On a side note: I'm also showing sluggish yeast after freezing. I'm seeing an easy 24-48 lag from pitching in starter to it actually starting. I suspect that freezing will affect different strains differently. I had a US05 come back after about 24 hours (frozen 2 months), but my 3787 trappish high gravity has taken 2-3 days just to awaken and it was only frozen for about a week.
Cheers!
It's all in the math
I have 50ml vials.
I create a 50% glycerin solution because it's easy to mix.
I want 15% glycerin concentration in my vials.
15% of 50ml is 7.5 ml of GLYCERIN.
Double that to 15ml because my stock solution is only 50% glycerin.
(50ml * 0.15 / 0.50 = 15 ml of 50% glycerin solution to 35 ml yeast slurry)
I can then fill the vial up with any amount of anything (except glycerin) and maintain a 15% glycerin concentration in my vial. Regardless of how thick/thin my yeast slurry is, as long as I fill up to 50ml the 15% glycerin concentration is maintained. If I'm a little short in one vial then I top up with sterile water to the 50ml mark - no problem.
The hard/questionable part is estimating how many billion cells you have put into each vial AND it's viability once thaw.
On a side note: I'm also showing sluggish yeast after freezing. I'm seeing an easy 24-48 lag from pitching in starter to it actually starting. I suspect that freezing will affect different strains differently. I had a US05 come back after about 24 hours (frozen 2 months), but my 3787 trappish high gravity has taken 2-3 days just to awaken and it was only frozen for about a week.
Cheers!
stpug: Your calculation summary is right on. On the other issue, I have belgian and american ale yeast as well as several lager yeasts frozen. I have used most of them after many months in the freezer but I am freezing at -80C in a non-frost free refrigerator. I always grow a starter. I see increase in cells in my starter after 4-8 hours. When I grow an overnight starter I have airlock activity within a couple hours of pitching. I don't have any experience with the frost free freezer method but would be interested in hearing what others have seen.
stpug: Your calculation summary is right on. On the other issue, I have belgian and american ale yeast as well as several lager yeasts frozen. I have used most of them after many months in the freezer but I am freezing at -80C in a non-frost free refrigerator. I always grow a starter. I see increase in cells in my starter after 4-8 hours. When I grow an overnight starter I have airlock activity within a couple hours of pitching. I don't have any experience with the frost free freezer method but would be interested in hearing what others have seen.
beernardo
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Hey, getting back into brewing and I'll definitely be trying this. One dumb-guy question here, perhaps. Is there any reason why I wouldn't use cheap vodka instead of isopropyl alcohol to slow down freezing? Seems like it would do the same thing that you suggest, wouldn't it? Also, it's cheap, easy to locate and replace, food-safe in case of an accident, and comes in a convenient container with a handle.
@beernardo I see no reason why Vodka wouldn't be perfectly fine. It won't freeze at typical home freezer temperatures and you can drink it when your done Here's an idea. Partially fill a plastic bag and then just push your tubes or bottles into a container with the plastic bag so it conforms to the tubes keeping them cold and your vodka pure.
Here's an idea. Partially fill a plastic bag and then just push your tubes or bottles into a container with the plastic bag so it conforms to the tubes keeping them cold and your vodka pure.tonyfelloni
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Hi guys.
This is some interesting work so thanks for sharing. I do have one question though. You say you have to wait extra time for the yeast to become active. Has anyone tried a control test where there control wort is exposed to the same environment as the wort with freshly thawed pitched yeast (no yeast added to the control)? I see fermentation start in wort after 2 to 3 days that I have pitched no yeast in, ie it has come from the surrounding air.
This is some interesting work so thanks for sharing. I do have one question though. You say you have to wait extra time for the yeast to become active. Has anyone tried a control test where there control wort is exposed to the same environment as the wort with freshly thawed pitched yeast (no yeast added to the control)? I see fermentation start in wort after 2 to 3 days that I have pitched no yeast in, ie it has come from the surrounding air.
tonyfelloni, If your wort undergoes fermentation after 2-3 days you have contamination. That is not entirely unexpected but suggests you aren't being as clean as you should be. The contaminants are probably kept at bay by the ongoing yeast fermentation if it gets a good start before they do. When I say the lag is longer before fermentation starts, I am talking about hours not days. Usually it is well underway within 12 hours but sometimes it takes somewhat more. Never 2 days. When I do a big fresh starter using that yeast it gets my airlock going within 2-4 hours. I don't think the side by side is necessary.
squibload
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Great thread!
I used to have 10 or 12 test tubes in the freezer at any given time, used agar (kinda like jello?) and inoculated it with yeast cells. Just a dab in each tube but enough to make a starter. Stretched one pack of liquid yeast to many batches.
Already read the yeast washing thread mentioned above, would make for a higher concentration to work with.
A discussion on yeast starters for bread got me here, thinking of a frozen "backup" in case the starter got compromised. I'll explore the site, was thinking about brewing again. My last batch was a mead, fermented for 10 months and I couldn't wait, bottled it up.
Could have gone longer but was tasty.
I used to have 10 or 12 test tubes in the freezer at any given time, used agar (kinda like jello?) and inoculated it with yeast cells. Just a dab in each tube but enough to make a starter. Stretched one pack of liquid yeast to many batches.
Already read the yeast washing thread mentioned above, would make for a higher concentration to work with.
A discussion on yeast starters for bread got me here, thinking of a frozen "backup" in case the starter got compromised. I'll explore the site, was thinking about brewing again. My last batch was a mead, fermented for 10 months and I couldn't wait, bottled it up.
Could have gone longer but was tasty.
Reviving an older thread here but this looks very interesting. A few questions.
1) If I were to purchase new vials of yeast and separate into multiples, these would all be considered 1st gen, correct?
2) Could you use the RV antifreeze as a medium to slow the freezing process of the yeast.
1) If I were to purchase new vials of yeast and separate into multiples, these would all be considered 1st gen, correct?
2) Could you use the RV antifreeze as a medium to slow the freezing process of the yeast.
radiogorillaz
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One could also probably use breast milk tubes. They have a flat wider base and will stand on their own. They are also intended to be frozen. Like these: http://www.amazon.com/gp/aw/d/B0009XBX4K/ref=mp_s_a_1_1?qid=1422290329&sr=8-1&pi=AC_SY200_QL40&dpPl=1&dpID=313lrM9glnL&ref=plSrch
MuddyCat
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Thanks BrewiTT for a very detailed and educational article. It, by far, has been the best one that I have ran across - and thanks to the folks who have posted comments as well. ALL great stuff, that is for sure!!
I am going to take a stab at freezing yeast, I've washed some leftover yeast from a couple 5 gallon carboy beer fermentations (primary fermenter) with good success thanks to Bernie's article which y'all can find here: https://www.homebrewtalk.com/f163/yeast-washing-illustrated-41768/
I was wondering if I could get away from using rubbing alcohol or gel packs as insulators by using an insulated lunch bag. So my idea is to mix the yeast and the glycerine into 50ml centrifuge tubes as indicated in Brewitt's "Freezing Yeast" article, place them in a test tube rack, and then cool the vials in the fridge overnight.
The next day, take the test tubes and the rack that holds them and place them directly into the insulated lunch bag, zip it up, and place it into the freezer. If this works the lunch bag will hold 2 test tube racks, with each test tube rack holding 24 - 50ml tubes, which would be pretty nice!
Do you think the insulated lunch bag would provide adequate protection against the defrost cycle on my "home grade frost-free" freezer? I also have a solution for handling storing of the yeast in alcohol, but if you think the freezer bag would insulate well enough - that would be a better solution as it simplifies the storage process. What do you think? Would it be recommended / suggested to place ice packs in the lunch bag? I guess it wouldn't hurt and it would help keep the yeast cold for a longer period of time in the event of a power outage.
Oh this may be handy for some folks who are looking for supplies. It is a link to a list of items that I am planning on using to freeze yeast:
http://www.amazon.com/gp/registry/wishlist/110QWN1UI24JN/ref=cm_wl_rlist_go_o?
The plastic boxes are the closest thing I could find to accommodate the test tube rack dimensions. Which if the lunch bag idea works, I won't need the plastic boxes.
Thoughts?
Thanks !!
I am going to take a stab at freezing yeast, I've washed some leftover yeast from a couple 5 gallon carboy beer fermentations (primary fermenter) with good success thanks to Bernie's article which y'all can find here: https://www.homebrewtalk.com/f163/yeast-washing-illustrated-41768/
I was wondering if I could get away from using rubbing alcohol or gel packs as insulators by using an insulated lunch bag. So my idea is to mix the yeast and the glycerine into 50ml centrifuge tubes as indicated in Brewitt's "Freezing Yeast" article, place them in a test tube rack, and then cool the vials in the fridge overnight.
The next day, take the test tubes and the rack that holds them and place them directly into the insulated lunch bag, zip it up, and place it into the freezer. If this works the lunch bag will hold 2 test tube racks, with each test tube rack holding 24 - 50ml tubes, which would be pretty nice!
Do you think the insulated lunch bag would provide adequate protection against the defrost cycle on my "home grade frost-free" freezer? I also have a solution for handling storing of the yeast in alcohol, but if you think the freezer bag would insulate well enough - that would be a better solution as it simplifies the storage process. What do you think? Would it be recommended / suggested to place ice packs in the lunch bag? I guess it wouldn't hurt and it would help keep the yeast cold for a longer period of time in the event of a power outage.
Oh this may be handy for some folks who are looking for supplies. It is a link to a list of items that I am planning on using to freeze yeast:
http://www.amazon.com/gp/registry/wishlist/110QWN1UI24JN/ref=cm_wl_rlist_go_o?
The plastic boxes are the closest thing I could find to accommodate the test tube rack dimensions. Which if the lunch bag idea works, I won't need the plastic boxes.
Thoughts?
Thanks !!
peterlonz
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I have missed something obviously.
What on earth is the isopropyl alcohol used for?
Its not low cost & its toxic.
I'd appreciate a bit of help here.
Thanks
What on earth is the isopropyl alcohol used for?
Its not low cost & its toxic.
I'd appreciate a bit of help here.
Thanks
Isopropyl alcohol is rubbing alcohol. You can buy it at the drug store or Target or many other places. It is not being used in the yeast slurry, it is being used as a non-freezing solution to keep the temperature more constant during the freeze-thaw cycles of a self-defrosting freezer. The bath is just made up once and reused. I prefer the gel from freezer gel packs but the alcohol will work.
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