Guide to Making a Frozen Yeast Bank

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Nightbiker said:
Thanks! I'll give 'em a look-see. I'm assuming that borosilicate glass (it says that the cap is even autoclavable -which is excellent) is also freezer safe?

Yes...
 
This may be helpful...

I recently bought 4 of these: http://www.midwayusa.com/product/15...ox-10-12-gauge-2-3-4-3-3-1-2-25-round-plastic in the smoke gray to hold the Cynmar 16ml vials.

Without any modding these are worthless. The vials can be forced into them but when the temp drops they are impossible to remove. The reason for that is inside each square is a tapered ridge, I assume it is used to keep the shotgun shell snug.

A quick trip to the craft store (I went to Hobby Lobby) and I bought a mixed pack of Exacto knife blades that had a thin, flat scrapper style blade. It was tedious but I scraped everyone of the ridges off and now the holder/box works like a charm. It was not hard to do but very tedious and a little messy with all the plastic bits.

Now they are all together and MUCH harder to knock over. Each box holds 25 vials I normally make 10-12 vials of each strain I bank so it works out well for me. :)
 
Uhm, I know I missed it, but where are you getting those vials? I'd love to have a batch of them -would really prefer larger ones (16 is such a small amount) -I've BEEN using sterile sample containers like we use in the hospital (110ml). I've never tried to harvest from a commercial bottle -and didn't know Bells bottle-conditioned. I love their brown ale -had it out of state once. Will have to see if I can find it and harvest some.
-thanks. Those vials look GREAT (love the tan look of yeast!)

Try out these: http://www.amazon.com/gp/product/B005Z4QSOS/ref=oh_o01_s00_i00_details

15mL conical centrifuge tubes pack of 100 for $16. They are autoclavable and can handle -80C temps fine, I use them in the lab all the time. If you really want you can wash and re-use them.
 
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Thanks Postal -those look like a really nice deal. I did find a container at the hospital (where I work) -they are sterile sample containers -measuring 110ml with a locking, sealling cap -made of plastic -and I have a couple of yeast samples freezing as we speak. They cost more than the tubes from Amazon tho (about a quarter per unit -but then again, they are also quite a bit larger) -I'll keep using the larger sample container, but I think it would be smart to duplicate my efforts with the vials you listed (in a different freezer -just to be safe. Probably more effort than it's worth, but you know how it is :)) We lost our big stand-up deep freezer -and along with it, all of my fresh frozen home-grown hops (cascades and mt. hood for the most part) -didn't notice it until it had completely defrosted, and the hops had turned a nasty shade of brown, and I won't even bother describing the status of my stockpile of venison and beefsteak.
Needless to say, I had to have quite a few consolation homebrews to get past that :(
 
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I'm not sure why people freeze the yeast. The liquid yeast isn't frozen when you get it and has a shelf life of a few months. Seems like with some planning you could just use one strain of yeast for several beers in a row within a couple of months...? Are people saving the yeast for long periods of time?
 
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liebertron said:
I'm not sure why people freeze the yeast. The liquid yeast isn't frozen when you get it and has a shelf life of a few months. Seems like with some planning you could just use one strain of yeast for several beers in a row within a couple of months...? Are people saving the yeast for long periods of time?

Some of the yeast I have frozen, you cannot buy from the store. So, once I got it cultured up from the bottle I freeze a whole bunch and can pull it out next year or later and make a starter.

Plus the viability of yeast simply washed and stored in the fridge is not good at all.... Maybe for 2 weeks, after that you definitely need a starter anyway. The yeast you buy from white labs is not even close to the same quality that you have coming out of your fermentor. Dead yeast, hops, alcohol and bacteria all deteriorate a washed yeast slurry very quick.

I do wash yeast, and try to plan it out so I am brewing the same day and pitch a good healthy yeast. But if it doesn't work out, I know I can make a starter from the greatly more sanitary vials I have in the freezer. Not to mention you can buy a vial of American ale yeast, let's say, grow up a starter in sterilized wort then pull out a couple mL for freezing and pitch the rest. If you save 5 vials, you just cut your $7.50 yeast cost into 6 batches... So it saves money in the long run. The equipment is extra, and you can buy a lot of yeast for the money, but a lot of us have the stuff already.

With some extra equipment and little planning, I think I have bought two vials of white labs in the past few years. If I had not done this, I'd have spent $150 just on yeast alone. I had all the equipment, and now I most of my batches are under $20, $25 for this big ones.

It's not for everyone and it is definitely not necessary, but it works for a lot of us.
 
I thought about using those, but I've seen a lot of those plastic centrifuge tubes melt or deform in an autoclave.

I've never seen those tubes melt. They go through the autoclave at my work fine which runs at a higher temperature than my pressure cooker can attain. Just make sure to not screw them tightly or they will explode or collapse during cooling.
 
I've never seen those tubes melt. They go through the autoclave at my work fine which runs at a higher temperature than my pressure cooker can attain. Just make sure to not screw them tightly or they will explode or collapse during cooling.

I should clarify, I have seen the 50 mL disposable centrifuge tubes melt a bit in a lab autoclave....since then I assume they might all melt. It wasn't exploded or crushed, the part of the tube above the liquid inside was melted and sloughed to one side.

I have some here at work that are 15mL...I'll throw some in my pressure canner and see if they make it through.
 
I am not sure if anyone has trouble resuspending their vials, but I did. It took more than 10min of shaking to get it halfway their. So adequate a vortex mixer using an orbital sander.

First I tried just the sander alone, which worked so-so. I had to hold the vial pretty hard against the sander and tilt it just a bit to get a vortex. Next, I placed the silicone jar opener on the sander. This worked much better, but the vial ate right through the silicone because of the pressure.

I then took the four srews out holding the sander platform on the bearing. I cut some plywood to the size, drilled holes for longer screws in the factory hole positions, drilled holes for the vials... And used the short factory screws as anchor points for the rubber band stabilizers.

Before:
2012-02-03_17-32-08_644.jpg


Installed:
2012-02-03_17-33-09_150.jpg


30 seconds after:
2012-02-03_17-36-05_636.jpg



She ain't gonna win a beauty pageant, but she sure mixes year vials well. :)
 
I should clarify, I have seen the 50 mL disposable centrifuge tubes melt a bit in a lab autoclave....since then I assume they might all melt. It wasn't exploded or crushed, the part of the tube above the liquid inside was melted and sloughed to one side.

I have some here at work that are 15mL...I'll throw some in my pressure canner and see if they make it through.

Weird. I autoclave the 50mL tubes after they have been used too since we get them already sterile.

I just used my pressure cooker on about 50 of the 15mL tubes with 3-4mL of glycerol and they all survived except one which I closed too tightly and it collapsed when it cooled.

Also, do you have problems with the vials cracking if you fill them all the way up and then freeze? Seems the ice would expand and break the glass.
 
never had a problem getting my yeast back into suspension. When I take one from the frozen 'bank', I put it in the fridge a day before use, then on the day I'm going to use it (to make the starter) I take it out of the fridge so it achieves room temperature while I make the starter wort, then by the time my wort is cooled, the vial is 'warm' and a brief shake puts everything where it needs to be.
Sometimes (if I have a large enough sample -I use a smaller test-tube-like plastic vial for long-term samples, but also store a larger amount in a sterile sample container (from my work place) -it is a 120ml container that seals and seems to survive freezing with no ill effects -when I get 'em, they're sterile as long as the paper tape seal hasn't been broken) -these are USUALLY used for urine samples (either drug testing or to obtain a culture) -so in a way, I'm kind of using them for PRE-urine samples (grin). Anyway, I've been just defrosting THOSE like above, and pitching it directly to the wort with good results.
 
postal_penguin said:
Also, do you have problems with the vials cracking if you fill them all the way up and then freeze? Seems the ice would expand and break the glass.

No breaking yet. Most of the vials are not even frozen. Some are a little slushy, but still air travels through if I tilt them.



Nightbiker said:
never had a problem getting my yeast back into suspension.

I think mine just lack enough headspace to get a good shake going on. Plus, my yeast layer is 3/4" thick when it settles out. They definitely need more agitation than I can manually provide.
 
I notice that after a day or two in the freezer, that my samples begin to separate (30% glycerin, 60% yeast, 10% headspace)...this normal?
 
bjohnson29 said:
I notice that after a day or two in the freezer, that my samples begin to separate (30% glycerin, 60% yeast, 10% headspace)...this normal?

Normal. The glycerin probably won't freeze, so separation is inevitable. The yeast will already be dormant as long as you store them in the fridge for a day or two before putting them in the freezer.
 
The 15 mil falcon brand don't fully melt but they do lose their shape in our autoclave.

As far as the yeast slurry none of mine settle out in layers. Why are they not freezing in a few hours time? You only need 20-25% glycerol and mine always freeze fine. The yeast may start to settle but 25% glycerol should stay as a homogeneous mixture
 
mthompson said:
Normal. The glycerin probably won't freeze, so separation is inevitable. The yeast will already be dormant as long as you store them in the fridge for a day or two before putting them in the freezer.

If this is the case you have a problem. Glycerol is used to cryoprotect the yeast so if your glycerol is not mixed homogeneously then the cells are not protected. Are you sure it is not just liquid that became visible when the yeast started settling?
 
ekjohns said:
If this is the case you have a problem. Glycerol is used to cryoprotect the yeast so if your glycerol is not mixed homogeneously then the cells are not protected. Are you sure it is not just liquid that became visible when the yeast started settling?

Not a problem at all, actually. Since the mixture is not frozen, there is no crystallization and no cell damage. Science doesn't fully understand how the mixture of water and glycerol works, but different mixes freeze at different points. Pure glycerol freezes at 17°C, pure water at 0°C.... But some mixtures of the two can go down to -47°C.

Pretty interesting chemistry in those hydrogen bonds: data

I do 50/50 distilled water and glycerol. 8mL glycerol mix + 8mL thick settled slurry. Since you don't count the yeast cells in the total mixture because they are merely suspended solids, I figure my final mixture is somewhere around 40-45% glycerin.
 
First I agree with the above that if it is not frozen then don't worry, but I would like to point out a home freezer should be around -18C (0.4F). Freezing point of 30% glycerol is -9.5C so your tube should be frozen. Check your freezer cause it may be too warm (assuming the addition of yeast and wort does not raise the freezing point... science is a fickle b1tch).
% glycerol by volume / freezing temp (C)
10% -1.6C
20% -4.8C
30% -9.5C
40% -15.5C
50% -22C
60% -36.6C
70% -37.8C
80% -19.2C
90% -1.6C
100% 17C
 
Thank you to OP (FlyGuy) and to everyone that contributed in this great thread. I ordered vials today, already had everything else. I'm jumping on the frozen yeast bank band wagon. :mug:
 
Freezing point of 30% glycerol is -9.5C so your tube should be frozen. Check your freezer cause it may be too warm (assuming the addition of yeast and wort does not raise the freezing point... science is a fickle b1tch).
% glycerol by volume / freezing temp (C)
10% -1.6C
20% -4.8C
30% -9.5C
40% -15.5C
50% -22C
60% -36.6C
70% -37.8C
80% -19.2C
90% -1.6C
100% 17C

The slurry I add is the consistency of yogurt because I decant all the liquid off and use a sterile 60mL syringe to suck it up and transfer. I'd guess it is at most 10% liquid:90% solids. That's why I figure my final concentration is in the 45% range, ignoring the weight of the yeast. My freezer is at -20ºC (higher in the pic because I had to wipe the frost off it, which meant touching it). I checked my vials and only two were semi-frozen. Since my vials are mostly still liquid at -20ºC, this is a second confirmation of that concentration. I also use distilled water, which has no other dissolved minerals to get in the way of the glycerol-water interaction.
2012-02-16_08-50-04_124.jpg


I am still not convinced that if they were frozen it would matter if the yeast was suspended or not. The yeast are surrounded by the same mixture, if they are all over the place or clumped together. Again, science doesn't know why it works, just that it does work.

:mug:
 
I agree that if it is just yeast that has settled out they are still surrounded by the mix of water/glycerol. My concern was that the post said there was a layer of glycerol not "liquid" . It could just be a problem of how I read it to be that the glycerol was separating from the water, not just the yeast settling.
 
Ok, you guys are really starting to make my head hurt. NOW I'm confused. I just received my order of 1gal (approx as the stuff is sold by weight, not volume) of 'pure' glycerin, I need to know how to cut it -should I use a 50/50 or 30/70 (which I believe is what is usually sold with yeast bank kits) -I don't have a lab-grade freezer, just a stand-up deep freezer that usually keeps Venison, hops and my yeast bank (wrapped in freezer packs to stabilize the temp since its a frost-free freezer). I'm not likely to GET a lab-grade freezer in the near future (especially as times are 'tight' financially, like it is with everyone else). So can I get a more-or-less concensus on the cut of glycerine to distilled water? IF not, what makes the 30/70 better than the thicker 50/50 (or worse)? My current samples (where I used the store-bought mix that came with my yeast bank kit) are frozen solid -which is what I assumed was the goal.
 
I use .25 ml glycerol and add 1 ml of my yeast starter at high krausen. This cuts glycerol to about 20%. Store in my deep freeze at about 5°F. Have never had a problem with resurrecting any of my vials. Just followed advice of a grad student workin on yeast.
 
I use .25 ml glycerol and add 1 ml of my yeast starter at high krausen. This cuts glycerol to about 20%. Store in my deep freeze at about 5°F. Have never had a problem with resurrecting any of my vials. Just followed advice of a grad student workin on yeast.

I wish to cut the glycol and then freeze it for future use (will split the mix down into a number of vials to use later, to avoid having to defrost and then re-freeze large amounts at a time. SO if I read you correctly, I'd be OK with cutting it 30/70 (70% distilled water to 30% glycerine), right? Overall, I'm probably putting more stress into this than I need to, but hey, gotta keep the yeast healthy if you want to make good beer. And re-using yeast makes for a signifficant cut in the cost of brewing.
 
30/70 is perfect. Really it can be anywhere between 15-30% and everything will be fine. what I do is take 300ul of glycerol and add 700ul of yeast right from a white labs tube then freeze. The glycerol is thick so I never get all 300ul so mine are probably around 25% glycerol which is in the 15-30% range. Storing in a house hold freezer is fine. The only concern would be when the freezer goes through a defrost cycle (ignore this if your fridge does not). If your freezer does defrost put the vials in a small cooler with ice packs to prevent them from thawing.

short version - aim for 30% and anything around there would work fine.
 
ul ? Yeah, I keep 'em wrapped up in freezer packs and have had good results so far. Getting ready to run a large batch (double batch) of brown ale, and will harvest that yeast once fermentation is complete. Mmmmm brown ale.... anyway! thanks for putting my mind at ease. I'll split this batch of glycerine up and freeze the bulk of it, leaving smaller amounts for ease of use (much easier and less hassle to defrost a small amount of the solution -since I never really know if this stuff will separate out, I always give a good shake to the solution before adding yeast.
 
Excited! I just bought some vials and misc supplies for this (and some plating and slanting).

I didn't read the whole 64 pages to see if this has been asked before, but I assume this method works just fine for Brett? And I think I read somewhere the Lacto freezes well also?
 
My bank is officially started! I took two samples direct from a new vial of WLP002 and made two 15ml frozen samples. Then I pitched the rest into a starter and have it cultured up to 1L. Tomorrow I will take 10 samples from this first gen starter and freeze them. Then I'll pitch the rest of the starter into an English Porter I'm brewing tomorrow as well.
That will be 12 vials in all of WLP002. After that I work on the WLP001, WLP005 and WLP090 already chillin' in the fridge :cool:

Cheers! :mug:
 
I ordered my glassware yesterday, have a DIY stir plate in progress, and I just ordered 5 strains of liquid yeast today. I noticed this thread started in '07 and I have a question for those of you who have been doing this awhile...

Do you have any data on the percentage of your frozen yeast cultures that are viable after 1, 3, 6 months, 1 year, etc..? The reason I ask is I'm trying to decide how many separate vials to make of a particular yeast specimen so that I have a steady supply of that strain.

Thank you for all the great info;
Steve
 
FYI, you can buy glycerin locally. I don't bank anymore (not pure yeast, anyway) but I used to get glycerin at Walmart, in the pharma aisles. I probably posted this info about 100 pages back in this thread, but thought I'd throw it out there again.
 
You did say that, and I remembered finally one day at Walmart. :rockin:

FYI, you can buy glycerin locally. I don't bank anymore (not pure yeast, anyway) but I used to get glycerin at Walmart, in the pharma aisles. I probably posted this info about 100 pages back in this thread, but thought I'd throw it out there again.
 
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