Yeast storage question

[dunbruha]

I am getting ready to start using liquid yeast, and I have been doing a lot of reading about starters and yeast storage methods. I am a bacteriologist by training, but I am discovering that yeasts grow quite differently than bacteria… I’d appreciate it if folks would comment on what I am planning, as I haven’t really seen anything quite like what I am thinking…

My goal is to develop a system that produces a consistent supply of yeast for the minimal cost, without having to deal with agar slants (and having to perform multiple step-ups).


Yeast storage

1. Open a fresh tube of yeast. I’m going to assume 75% viability, or 75B cells total. I’m guessing there is about 30-35 ml of slurry in the tube.

2. Using a sanitized pipet, add 3 ml of the yeast slurry to vials containing 3 ml of sterile 20% glycerin. Put in fridge for 24h, and then freeze. I’m estimating I can get 10 vials from each tube, with each vial having 7B cells.


Starter preparation

1. Thaw a vial to room temperature, then add to a sanitized flask containing 1 quart of sterile wort (1.036) plus some yeast nutrient. Incubate on a stir plate at 77*F for 24h.

2. Add this culture to 1.5L of wort (1.036, or higher for high-gravity beers). Incubate again on stir plate @ 77*F for 24h. Cold crash, then pitch. I am guesstimating that I would have approximately 300B cells at this point (based on the Braukaiser - Stir plate option in the Brewer’s Friend yeast calculator).


What do you think? Do you see any potential problems, or any improvements? Thanks in advance—I have learned so much from reading HBT!

 

[fimpster]

My favorite yeast calc is THIS ONE. It calculates 348 bn cells using Kai's (brukaiser) equation. Either way you'll likely have plenty of cells for even moderately high gravities.

This sounds really interesting, looking forward to how this turns out.

Edit: I just ran your numbers through Brewers Friend and got 367 bn.

 

[PoppinCaps]

I would (and do) make a 800ml starter with the vial, cold crash, decant and aliquot to 8 conicals. Double that volume with 20% glycerine as you do and freeze in an ethanol bath.

This way you can thaw the conical, add to whatever size starter you want and be ready to cold crash in about 24 hrs for the brew. The difference between bacterial culture and yeast is great, as you said, and I think one of those differences is inoculation density. Bacterial cultures will take off with very little cells, but I'd be cautious in diluting yeast cells 1:300 from their peak density. I like a 1:10-1:20 dilution for quick guaranteed starter prep.

Just my take.

Edit: Nevermind, I saw that you're taking 3ml of the packed slurry, not a decant. Still, keep in mind the overall starting density of the cells in the starter, and try to keep it similar to the inoculation target of the beer you're brewing rather than a strict 3ml per starter.

 

[dunbruha]

I would (and do) make a 800ml starter with the vial, cold crash, decant and aliquot to 8 conicals. Double that volume with 20% glycerine as you do and freeze in an ethanol bath.

Thanks for your reply. Yes, this is my other favorite alternative. This one is quicker from freezer to pitching, but it does require a starter from the original, and I was worried that it might become contaminated, and thus all of my stocks would be bad...

I hadn't seen any mention of going straight from the original tube to freezing, and was wondering if it would work, or if there was some issues with it that I hadn't thought of.

 

[PoppinCaps]

Got it. There's no problem that I can see about freezing straight from the vial, the only thing may be the unknown viability of the stock, which would leave an unknown before brew day.

 

[dunbruha]

I would (and do) make a 800ml starter with the vial, cold crash, decant and aliquot to 8 conicals. Double that volume with 20% glycerine as you do and freeze in an ethanol bath.

So after you decant, approximately how much volume are you adding to each conical?

 

[PoppinCaps]

It's usually between 10-15ml. The volume per conical doesn't matter much to me, but rather the number of cells in each. I shoot for 5-8 conicals per 800ml decanted starter, to seed new 800ml starters after thawing, a little heavy to account for some loss of viability after freeze/thaw.

 

[dunbruha]

It's usually between 10-15ml. The volume per conical doesn't matter much to me, but rather the number of cells in each. I shoot for 5-8 conicals per 800ml decanted starter, to seed new 800ml starters after thawing, a little heavy to account for some loss of viability after freeze/thaw.

Thanks. I appreciate your answers. You say that you shoot for 5-8 conicals--is the variability based on the age of the original yeast tube, or do you do some kind of cell count after incubation?

 

[BobBailey]

Not very scientific, but really simple. Make a starter from the vial of yeast that is adequate for the beer you're brewing, plus an extra 100B cells. Refill the vial with the 100B cells and re-date it. Treat it as you would the original vial. Now you have a fresh vial and a proper pitch. This can be repeated 5 to 10 times.

 

[PoppinCaps]

Thanks. I appreciate your answers. You say that you shoot for 5-8 conicals--is the variability based on the age of the original yeast tube, or do you do some kind of cell count after incubation?

Depends on the starting volume mainly, but I usually stick with 5 tubes per 800ml starter. I have done cell counts when I restocked and froze my whole bank, and the final densities are very different depending on the strain. I guess that's why I stick with a static volume dilution for stocks, rather than number of cells; some strains just like their room. Seems to work. Just unthawed a conical of PacMan yesterday, it's already done now and cold crashing tonight.