Frozen Yeast Bank

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Postby Legman » Sun Jan 25, 2009 7:58 am

This is from the very first vials I made, no yeast washing and from the primary. I'll be making a starter and harvesting from that from now on. It's much cleaner and probably contains more viable yeast. I was impatient and couldn't wait to try it and that was just what I had on hand at the time. The other vials look like what you would buy at your HBS. But this kinda gives you an idea. Shake them well before you freeze them.

http://farm4.static.flickr.com/3122/322 ... 83.jpg?v=0
http://farm4.static.flickr.com/3338/322 ... a6.jpg?v=0
http://farm4.static.flickr.com/3442/322 ... d6.jpg?v=0
http://farm4.static.flickr.com/3378/322 ... fe.jpg?v=0
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RE Frozen Yeast Bank

Postby wottaguy » Sun Jan 25, 2009 11:17 am

Thanks for the pics Legman. I ended up ordering the "yeast bank" product from Country Wines and also got some extra vials. Here's the url in case anyone's interested:

http://www.countrywines.com/products.asp?category=150

I already have a couple of starters made and will try to post pics of my process when I harvest and freeze.

BTW...R U a race fan? Coming down for the 500??? Just wondering...

Thanks!

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Freezing

Postby slothrob » Sun Jan 25, 2009 11:22 am

I don't freeze yeast, but I freeze human cells, so take my advice with some skepticism.

The big problem with freezing cells is ice crystal formation. This problem is partially reduced by using glycerin, but not completely removed.

Cells are frozen in vials that are about 3/8" across. They can be longer but not fatter (the largest I know of are 5 mL). The reason is that the narrow diameter dissipates heat better. What happens is that when the fluid crystalizes a small amount of heat is given off. This causes the solution to thaw and refreeze. Each refreezing cycle kills more cells. The narrower tubes allow more of this heat to dissipate while fat tubes cause repeated micro-freeze-thaw cycles.

It's very similar to the reason you try to reduce the effect of defrost cycles by putting the vials in a secondary container with an ice pack.

What I'm trying to get at is that you may end up with as many viable cells from a small vial as a larger one. Unfortunately, cryovials are expensive. You might get better recovery from a thinner tube, like these 15 mL tubes, though.

By the way, it's best to freeze slowly, but get them across the freezing threshold quickly, then thaw quickly. So, refrigerate your yeast, glycerine and vials first, then stick your filled vials in something insulated, like a styrofoam box, to freeze. When you want to thaw them, take them from the freezer then whirl them around in warm water just until all the ice is gone, then keep them refrigerated until you pitch your starter, which should be sooner than later.

I'm glad what you're doing is working, but a few tweaks might help. You should probably pitch initially into a small volume of 1.020ish wort, to reduce shock to the roughed-up yeast, then step up to a full-sized starter once you get revived activity.
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tubes

Postby slothrob » Sun Jan 25, 2009 11:28 am

I think the tubes in wottaguy's kit are 5 mL tubes and are about the right diameter. They're a cheaper tube than the ones I use in the lab, so they're probably a decent choice for home.

I can't find a cheap non-industry source for them, though. Do they charge $1/tube? I think we pay about $0.10-0.20/tube, but we buy them 1000 at a time.
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Re: Freezing

Postby Legman » Sun Jan 25, 2009 11:42 am

OK Slothrob, I'm not gonna ask why you're freezing human cell down in your basement. (Trying to create the perfect woman) :shock:

Just kidding, I'm sure that's work related. You may be right about the tubes and freezing process. That may be something I look into later. Like everything we do in brewing, it's all a natural progression to improve. But for now, I'll keep as is and see what kind of results I keep getting. Thanks for the info though. That may be helpful in the future.

And I gotta ask......is it Slo-throb or Sloth-rob. Hehe. :lol:

Wottaguy, no I'm not a race fan. But I've been to several of them and had a good time. That's definitely a beer drinking sport!
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Re: Freezing

Postby wottaguy » Sun Jan 25, 2009 11:54 am

Legman wrote: And I gotta ask......is it Slo-throb or Sloth-rob. Hehe. :lol:



Ya know...i've been wondering about that same thing for the longest time now, but wasn't sure if i should ask!! LOL!! Sooo....which is it??

:D

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freezing

Postby slothrob » Sun Jan 25, 2009 1:47 pm

Well, it's both, isn't it? :wink:

If it's any help (perhaps if you were an English major or interested in surrealism...well, you asked...), it started out as a typo of Slothrop, about 15 years ago, and it's just been inconvenient to change it. My surname is way too common to ever be available as a login or email.

You can call me Eric.

Yeah, I do research in growing human cells to produce replacement organs for transplantation. A long time ago I took training in cell freezing at the ATCC, which is the worldwide center for storing frozen cell stocks, and I've frozen hundreds of cell strains over the years. My yeast experience is mostly limited to brewing, though. Yeast are a little more forgiving, when it comes to freezing, because the cells are smaller, but the general theory is the same.[/i]
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RE: Yeast Bank

Postby wottaguy » Fri Jan 30, 2009 8:45 am

Well guy's, I have officially started my yeast bank and harvested 3 vials of WLP051 Cali V Ale. I created a starter this past Monday and let it ferment out, then put in fridge to settle the yeast out. on Wed, I created more wort from DME and decanted the cooled off starter's clear liquid, and refilled it with the new cooled wort. This starter started fermenting in an hour of doing this and was very active. I let it go for 4 hours, then put it on my stir plate in a temp controlled fridge (65 deF) and let it ferment out. This morning i prepared and sanitized 3 vials with caps, a gravy baster and my working counter with starsan and filled the vials up with the very milky yeast starter, then returned the starter flask to the fridge and the stir plate. I labeled the vials and put them in a small beer sampling glass, (the kind you get at comps and festivals), and placed the caps on loosely, and will let them sit and settle over night.

Tomorrow, I'll take the settled vials out, and with a sanitized eye dropper, will suck most of the clear liquid out, then refill with my glycerin solution, wrap with paper towels, and place in my freezer for long time storage.

The rest of the starter will be used on Sunday for a Blond Ale.

Next up....Irish Ale Yeast..!

Can't wait to have a good collection of yeast on hand. It's going to take time, as I'm harvesting as I go from the starters that I make for my next brew, but should be worth it in the long run. Sorry...no pics...maybe next time.

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Yeast Bank

Postby Legman » Fri Jan 30, 2009 9:33 am

Wottaguy, how's that yeast bank kit you got? Does it says what the ingredience are? Intructions?
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Yeast Bank...

Postby wottaguy » Fri Jan 30, 2009 10:28 am

Legman....It came with full instructions, but doesn't say what the "yeast shield" is composed of. It also came with 5 test tubes, and I ordered 15 more of those to go with the kit. My plan is to make 3 tubes of yeast from my starters when I brew, to save for freezing. My only worry is about the hurricane seasons and the loss of power for any great length of time. We don't have a generator yet....but I can always run the yeast across the street to my neighbors place as they do have a generator.

I'll be freezing my first 3 tubes of yeast tonight or tomorrow morning.

Thanks for asking!

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Yeast Bank

Postby Legman » Fri Jan 30, 2009 1:17 pm

Cool. Good luck with it.
I don't know what your freezing instructions are, but I've read that you should pack them in a small cooler with ice packs to help with the automatic thaw cycle of the freezer. The constant thawing and freezing can kill the yeast.
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bak at ya...Yeast Bank

Postby wottaguy » Fri Jan 30, 2009 3:21 pm

yeah...i've read that too...so my plan is to wrap the vials in bubble wrap and keep them on the bottom of our freezer in a small plastic container. It will essentially work like the method that you described.

Thanks for your best wishes and of all the info you have passed along....i'll let ya know what happens after I thaw one out and make a starter with it....could be a while from now...lol

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Not much yeast for the bank

Postby shaggyt » Tue Feb 17, 2009 10:25 pm

I went out and got the yeast bank last week and began my starter (weaker than normal though). I filled my vials and pitched the starter last night. fermenter working throughout the day today and yeast has fallen in the vials.

So I'm about to draw off the liquid from the 5 vials of starter but I noticed there's only about a small amount of yeast in each vial. I shook the starter vigorously and used a baster to extract my samples for vials. I thought I got enough to fill more than just the very bottom of the vial.

My question to those who've harvested...what proportion (roughly) of yeast is the minimum needed to successfully harvest?

For my example, I'd say there's about 1-1.5ml of yeast at the bottom (just enough to fill the curved bottom of the vial).

Thoughts, advice, reassurance...it's all welcome at this point.

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Re:

Postby Legman » Wed Feb 18, 2009 5:52 am

Shaggyt, from what I've read, technically you don't need to have a large amount to propagate the yeast. But you'll probably have to do a step-up starter so it doesn't stress the yeast out.
I assume you're following the instructions that came with your yeast bank. If that it true and from what you described, what you collected per vial seems about right. You're not going to end up with a vial full.

But if you think you need to collect more per vial, make a regular starter, let it ferment to completion and settle to the bottom. Then siphon off most of the liquid, then stir it up and fill you vials. I too was concerned about the amount collected, so I've been trying to get as much as I can in a vial. Might be overkill, but it makes me feel better anyways. I've got vials just like White Labs comes in and I'm maybe getting about 2/3 as much yeast in a vial as if you were to buy one.

There's going to be some trial and error in this process. So far I've got about 25 vials in the freezer. I'm not by any means an expert at this and I've only resurrected one vial so far....but it did work.

Good luck and keep us posted.
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RE: Not much yeast for the bank

Postby wottaguy » Thu Feb 19, 2009 10:08 am

ShaggyT...

I too have just began to harvest yeast and freezing samples for later use, and I am using the Yeast Bank as well. I also thought that the volume of yeast first collected from the starter didn't look like enough, so when the starter was spent, I put it in the fridge to cool down and settle the yeast, and once settled I took it out of the fridge to warm it back up, and made a second wort medium while it was warming up, and cooled the wort off to room temp and decanted the clear liquid from the starter and then put the new wort medium in the flask again to restart the yeast. I put it on my stir plate and within 2 hours it was fermenting again. I watched the starter and when I was satisfied that the starter was in full bloom, I took my baster out, sanitized it...decanted off the clear liquid from the vials and essentially top cropped the yeast from the starter and put it in the test tubes, and returned them back into the fridge. The next morning I was happy to see that I had a lot more yeast in the vial and very little trub from the starter too, and ended up with about 2 ml in each vial, which should be plenty to make a 500ml starter from. I then decanted the clear liquid (beer) from the vials and added the "freeze shield" , mixed them by gently shaking each vial until very well mixed then wrapped each vial in a paper towel, labeled them and put them in the freezer for later use.

Like Legman stated, we'll more than likely will have to step up the starter a couple of times before pitching in any brews, but the savings will be worth it!

I haven't resurrected any frozen yeast yet as I am building up my bank and have only 3 strains frozen so far.

Let us know how you make out with your yeast bank and I'll be doing the same.

Good luck!

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