Do you know how to make a yeast starter? Then why not farm yeast and freeze it?

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If you're going to use 15% glycerol then you'll want to let the cells settle out and decant as much of the liquid off the top as possible. If you're slurry is quite watery, then you'll want to increase the % glycerol.
 
Good to know that the solution with lower glycerine content give the best results.
This means cheaper freezing process and possibility to fill the vials with more yeast!:mug:
 
Forkhead said:
If you're going to use 15% glycerol then you'll want to let the cells settle out and decant as much of the liquid off the top as possible. If you're slurry is quite watery, then you'll want to increase the % glycerol.
Click to expand...

If the yeast are pourable it is probably best just to use a equal volume of 30% glycerol if you can.
 
Nice work Forkhead. It would be nice to see that repeated a couple times (not asking you to). I'm just curious as to why viability was better at -20 than -80 on the short term. Perhaps becasue the -80 aliquots froze faster?
 
My last statement about using 30% glycerol and an equal volume of cell slurry applies to -80 freezing. I forget that most of us don't have access to that sort of freezer (and believe me, they are very expensive). So, based upon Forkhead's results, it should be equal volumes of yeast slurry and 15% glycerol for freezing at -20. It is great that 7.5% glycerol provides a good protective effect.

With response to BBL_Brewer's comment, I think the error in the viability determinations depends upon how many colonies were counted in the plating test, which is certainly the most accurate determination. If it was not a large number then those numbers are within the error. For example, the -20 with 7.5% final glycerol is higher than the starting viability (doubtful that the freezing in glycerol revived dead yeast ;-). Forkhead, would you agree with that assessment of the viability numbers or do you know that the reliability is higher than I am suggesting. I would also point out that there will be some differences between yeast strains in cryosensitivity and such. These should probably be taken as relative numbers (i.e., best versus worst conditions rather than that a particular condition gives x % viability).

Forkhead, it will be interesting to see what the longevity of the frozen yeast is at -20. I imagine that leaving them in isopropanol would be good for storing in a frost free refrigerator. Even a large bottle or container with isopropanol containing baby food bottles or water tight plastic containers would work for that kind of setup and act as a barrier to thawing. The low glycerol and isopropanol might overcome the biggest barriers to long term yeast storage by home brewers.
 
I used one of my thermometers (sensor connected to the unit, sensor inside freezer, unit outside) to test both of my freezers. The one that holds hops doesn't get down to 0F. The one that's used for food gets below 0F. It actually gets down to about -10F and goes up to about -6F before cooling down again. Both are set to their max cooling settings (for the freezers). Need to see about getting, or making, a small 'cooler' box to put the vials into. Looks like I'll end up using the freezer that's part of the food fridge for yeast storage. Unless I can score a small freezer that will also get cold enough. Could use that for hops and yeast storage. :D
 
Brewitt said:
My last statement about using 30% glycerol and an equal volume of cell slurry applies to -80 freezing. I forget that most of us don't have access to that sort of freezer (and believe me, they are very expensive). So, based upon Forkhead's results, it should be equal volumes of yeast slurry and 15% glycerol for freezing at -20. It is great that 7.5% glycerol provides a good protective effect.
Click to expand...

When I did my freezing, I actually centrifuged the cells and removed the liquid before adding the glycerol solutions. As a result the yeast wasn't really a slurry and the glycerol wasn't diluted very much by residual water. If your yeast is actually a slurry then it might be better to use a higher concentration of glycerol (20-30%) to account for the dilution caused by the water in the slurry. However, i don't think we know the minimum amount of glycerol that's necessary. Maybe that should be tested. I hope that makes sense.

Brewitt said:
With response to BBL_Brewer's comment, I think the error in the viability determinations depends upon how many colonies were counted in the plating test, which is certainly the most accurate determination. If it was not a large number then those numbers are within the error. For example, the -20 with 7.5% final glycerol is higher than the starting viability (doubtful that the freezing in glycerol revived dead yeast ;-). Forkhead, would you agree with that assessment of the viability numbers or do you know that the reliability is higher than I am suggesting. I would also point out that there will be some differences between yeast strains in cryosensitivity and such. These should probably be taken as relative numbers (i.e., best versus worst conditions rather than that a particular condition gives x % viability).
Click to expand...

I agree that there's quite a bit of error in the measurement, which comes from a number of different sources that I don't really want to get into. I agree the point is not to be highly quantitative but rather to determine optimal conditions that gives very high viability. There's probably more than one way to do that. I think my results show no decrease in viability when using 15% at -20C vs storing at 4C. Increasing glycerol tended to have a negative impact on viability.

Brewitt said:
Forkhead, it will be interesting to see what the longevity of the frozen yeast is at -20. I imagine that leaving them in isopropanol would be good for storing in a frost free refrigerator. Even a large bottle or container with isopropanol containing baby food bottles or water tight plastic containers would work for that kind of setup and act as a barrier to thawing. The low glycerol and isopropanol might overcome the biggest barriers to long term yeast storage by home brewers.
Click to expand...

I'm guessing the long term viability will be better by moving the tubes from -20 to -80. I also don't think storing them in isopropanol is necessary after they've frozen unless you're using a conventional home freezer, which most homebrewers are. But if you have a non-defrosting freezer then the isopropanol won't matter after their frozen.
 
BBL_Brewer said:
Nice work Forkhead. It would be nice to see that repeated a couple times (not asking you to). I'm just curious as to why viability was better at -20 than -80 on the short term. Perhaps becasue the -80 aliquots froze faster?
Click to expand...

It's possible they froze faster. I'm thinking it'd be best to freeze them at -20C for a few days then move them to -80C. Alternatively, it's possible the colder temperature can more thoroughly freeze the tubes and may cause more ice crystals to form. That was actually predicted by that paper I cited a while back.
 
Golddiggie said:
I used one of my thermometers (sensor connected to the unit, sensor inside freezer, unit outside) to test both of my freezers. The one that holds hops doesn't get down to 0F. The one that's used for food gets below 0F. It actually gets down to about -10F and goes up to about -6F before cooling down again. Both are set to their max cooling settings (for the freezers). Need to see about getting, or making, a small 'cooler' box to put the vials into. Looks like I'll end up using the freezer that's part of the food fridge for yeast storage. Unless I can score a small freezer that will also get cold enough. Could use that for hops and yeast storage. :D
Click to expand...

Sound like your food fridge is perfect. Getting a cooler filled with isopropanol will help protect the cells from temperature fluctuations.
 
Forkhead said:
Sound like your food fridge is perfect. Getting a cooler filled with isopropanol will help protect the cells from temperature fluctuations.
Click to expand...

I made a 'cooler box' out of a cardboard box and styrofoam I had on hand. I can put 3, maybe 4, vials in there. Thinking about freezing the vials of yeast early in the coming week. That is, unless I get to it on Sunday.

Finished up a longer brew day today (make a big brew, biggest I've made so far) so I'm a bit beat.
 
That is good idea, especially with self defrosting freezers... luckily I got old fridge w/freezer that dont have that option, I hope it will live for a while..
 
Hey Forkhead. I've been running some numbers on YeastCalc and they're pretty close to the growth rates you reported a while back. You said you got ~250 billion cells from (3) 1.2 - 1.5 liter steps with a startinig cell count of 50 billion. Looks about right. Also, off to the side it shows you growth curves for White Labs, Wyeast, Mr. Malty, and YeastCalc.
 
Got mine from Amazon. $11.75+FREE shipping (Amazon Prime Membership :ban:)... Plus, the one on Amazon is 100% pure (DD is 99.7% pure), and it's Kosher USP food grade vegetable glycerin. :D
 
Golddiggie said:
Got mine from Amazon. $11.75+FREE shipping (Amazon Prime Membership :ban:)... Plus, the one on Amazon is 100% pure (DD is 99.7% pure), and it's Kosher USP food grade vegetable glycerin. :D
Click to expand...

The glycerine from amazon came from Essential Depot. If you look at the certificates of analysis from ED and DD, both products are vitually identical. In fact, the COA from DD shows it's product as being 99.9% pure whereas ED shows 99.84. What gets me is that DD is advertising their product as being made in the USA. However, they state that the source palm was grown in Mylasia and if you look at the COA, it clearly shows that the drums were shipped from Indonesia. In other words, not made in the USA. ED, on the other hand, plastered their generic logo on the COA and give no clues as to where the glycerine originated from. What this means is that the glycerine from amazon is no more pure or safe than that of DD. In fact, I would trust DD over ED since they are at least forthcoming about where the product came from and present their COA in it's original form from the lab.
 
If anybody needs a stir plate, I make them out of attractive cigar boxes, not cheap project boxes. I have some for 2 liter Erlenmeyer flasks that will make starter for a 10 gal batch using a 1 inch stir bar. I can also make a MONSTER stir plate using a 2 inch bar for a 5 liter flask. These stir plates use a pulse-width modulator for speed control. Those plans you see on the net have very limited speed control, mostly too fast. It's not necessary to beat your yeast like a red-headed stepchild. My stir plates let you control the speed to a nice gentle stir. Anybody interested, let me know for pictures and quotes.
 
tlb2525 said:
If anybody needs a stir plate, I make them out of attractive cigar boxes, not cheap project boxes. I have some for 2 liter Erlenmeyer flasks that will make starter for a 10 gal batch using a 1 inch stir bar. I can also make a MONSTER stir plate using a 2 inch bar for a 5 liter flask. These stir plates use a pulse-width modulator for speed control. Those plans you see on the net have very limited speed control, mostly too fast. It's not necessary to beat your yeast like a red-headed stepchild. My stir plates let you control the speed to a nice gentle stir. Anybody interested, let me know for pictures and quotes.
Click to expand...

You shouldn't be self-promoting without a vendor's license. And my stir plates work just fine, thanks.
 
Would anyone be offended if I consolidated the information gleaned from this thread and wrote up a Tutorial to post on the site. I think we have a critical mass of information that should be summarized. Credit where credit is due, of course. One of us should definitely do it.
 
Brewitt said:
Would anyone be offended if I consolidated the information gleaned from this thread and wrote up a Tutorial to post on the site. I think we have a critical mass of information that should be summarized. Credit where credit is due, of course. One of us should definitely do it.
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Shooting for a free membership? ;)

Doesn't bother me any.
 
Actually I wrote this when I got an email about an article on washing yeast. Then I noticed you could get a free membership. Might make it more worthwhile although I am not sure what a membership gives you. Do I get something good, besides a warm fuzzy feeling of membership?
 
I do want to give credit to those who have made substantial contributions to to protocol and I will cite a few papers I have seen. I would start with something of the format of BBL_Brewer's initial article but update it with the new cooling and thawing protocols, glycerol concentrations, and some of the viability information. It seems like Forkhead's isopropyl alcohol approach is a good one for evening out freezing and avoiding thawing. Another approach I have seen for avoiding thawing in frost free freezers is to use the gel packs that stay soft on freezing and surround your vessel or tube with them. I try and get as much of this stuff in as possible. Maybe take a shot at it this weekend. Although, being San Diego Beer Week it may be hard to get much done on it :D
 
OK, I decided to just go ahead and write an article last night. It is pretty close to finished including What, Why, Supplies, How, Resources, etc. I looked at the instructions for articles and they say I have to have at least three pictures. I took a picture of some empty tubes from White Labs yeast and some 50 ml screw top centrifuge that I use to freeze them. I also took one of a similar rack of frozen yeast I made from bottles of Pranqster (Belgian Golden Strong Ale from North Coast Brewing Co.) and Sculpin (a Ballast Point IPA). I could try to get together the whole set of supplies I suggested and take a picture. Those include a starter culture to be frozen, tubes, glycerine, water, isopropanol, a styrofoam container and/or a large container for freezing/storing tubes in isopropanol. Other suggestions for pictures?
 
Thanks. 16oz of glycerine or the big gallon jug? I'm having a hard time understanding the scope of this process.
 
theveganbrewer said:
Thanks. 16oz of glycerine or the big gallon jug? I'm having a hard time understanding the scope of this process.
Click to expand...

Unless you plan on freezing a LOT of yeast, I'd get the quart bottle first. You'll be diluting it, so that will go a long way. You can plan the next purchase according to how fast you use the first one up.
 
Golddiggie said:
Check out Amazon... I got some 50ml plastic vials/tubes from there. They also have glass ones, but getting them close to the same size isn't easy.
Click to expand...

Yes, 50 ml centrifuge tubes from Amazon are ideal. Here is the link for smaller quantities:

http://www.amazon.com/dp/B0018MQ9TK/?tag=skimlinks_replacement-20

I'm going to post my article soon. It has sources for glycerin and plastic tubes. Also a detailed discussion of the whole process summarized from this thread.
 
Last edited by a moderator:
I just submitted the article on Freezing Yeast. I don't know if its visible to just me or everyone but it is in the Technical Article section right now. I don't know what kind of file the moderator wants but I did it as a PDF file. Hopefully you all like it and agree with my distillation of the methods. In any case, what I proposed should work great and many successful modifications on the theme are possible.
 
So thanks to your links I found out those white lab tubes are called baby soda bottles, which opened up a ton of links to buy them.
 
Brewitt said:
I just submitted the article on Freezing Yeast. I don't know if its visible to just me or everyone but it is in the Technical Article section right now. I don't know what kind of file the moderator wants but I did it as a PDF file. Hopefully you all like it and agree with my distillation of the methods. In any case, what I proposed should work great and many successful modifications on the theme are possible.
Click to expand...

If you submitted it for a membership we can't see it, it's hidden.
 

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