How to build (step up) a starter

[The-Engineer-That-Brews]

I think building up a starter is easier than it sounds, but when I first did it I struggled to find a clear and well-informed set of instructions.

So I'm just posting this for future reference, from a microbiologist who's possibly a bit better-qualified than most on the subject:

The example I gave in class, and what I routinely do for lagers:​

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Target: 400 billion yeast cells (I just did this for a bock I'm brewing next week).​

The ideal dilution step for maximum growth in my hands is 1:10. I use a stir bar and stir plate as this is the only way to get maximal yeast growth.​

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1) 1 liter starter - 1.040 + 1 wyeast smack pack​

2) grow for 24, no more. This is key. After 24 hours the yeast will still start to go dormant, building up glycogen reserves. Starters should be "split" when the yeast cells are actively replicating. You should go from 100 billion cells to about 180 billion​

3) Dilute 1:10 into a new starter and grow for 24 hours. That means 100 mls into another 1 liter starter. This starter will go from 18 billion to another 100 billion.​

4) Cold crash the first starter, decant wort.​

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At this point you have close to 300 billion, but you need more.​

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6) Do a third starter from the second as in step number 3, grow for 24 hours.​

7) Combine starter #2 into #1, cold crash and decant.​

8) After the third starter finishes, you should have close to 400 billion. You can then combine all the starters and pitch into beer.​

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Keep in mind, this only works with a stirred starter. I believe intermittent oxygenation is not good enough, but rather the gas exchange with a stirred starter provides all the O2 for the yeast to grow. You may need multiple flasks, but not necessarily. You can always have another sanitized vessel on hand to collect the starters.​

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I particularly like the way that this technique sounds like it allows you to build a large number of cells without calling for a ridiculously large flask that barely fits on the stir-plate.

 

[RoomWithABrew]

If from wire loop from agar I start in sterile pot with 10ml at 1.030.
Increase by ten times so around 100ml, then one litre then I just scale it up to 2.5 litres.
If I'm starting with probable more cells just old then straight to 100ml, might then go to a few hundred then add a litre volume of wort aiming to have wort at 1.035 factoring in the dilution of existing wort.
I add nutrients and temp control as per sui generis recommends here.
https://suigenerisbrewing.com/index.php/2022/09/20/optimizing-yeast-starters/

Always use a stir bar, 3 litre flask base fits perfectly on my stirrer.

 

[The-Engineer-That-Brews]

@RoomWithABrew do you do your own yeast slopes?

 

[RoomWithABrew]

I was given a load of slopes. Most of my stock of yeasts is in sterile lab collection pots filled with a sample from starter or top cropped samples.
I've resuscitated these after 2 or 3 years in the fridge.

 

[The-Engineer-That-Brews]

I add nutrients and temp control as per sui generis recommends here.
https://suigenerisbrewing.com/index.php/2022/09/20/optimizing-yeast-starters/

Looks like a great resource

 

[Nicks90]

Ok maybe I am missing something really obvious here... But let's say you have the dregs of a bottle conditioned beer you want to harvest the yeast.

What's the difference between putting that 10ml of dregs in to
Scenario a)
1 - 100ml at 1.030
2 - pour the resultant yeast in to 1l at 1.030
3 - pour the resultant yeast in to 2.5l at 1.030
Scenario b)
1- 2.5l at 1.030 and leave it till it's finished

 

[The-Engineer-That-Brews]

Ok maybe I am missing something really obvious here... But let's say you have the dregs of a bottle conditioned beer you want to harvest the yeast.

What's the difference between putting that 10ml of dregs in to
Scenario a)
1 - 100ml at 1.030
2 - pour the resultant yeast in to 1l at 1.030
3 - pour the resultant yeast in to 2.5l at 1.030
Scenario b)
1- 2.5l at 1.030 and leave it till it's finished

Hi Nick!

The name of the game, as I understand it, is to persuade the yeast to reproduce rather than just convert sugar into alcohol.

So I think what happens is that when yeast is pitched into fresh wort, especially when there's plentiful oxygen available, is that it reproduces for about 24hrs and then switches over to just producing alcohol instead - probably as a means of suppressing the growth of competing organisms (with the fortunate side effect of getting humans drunk!!)

The idea of re-pitching it is therefore to keep it in "reproducing" mode rather than "producing alcohol" mode...

What the microbiologist says is:

The most important thing to do when stepping starters is not to add too much yeast to the next starter. This will inhibit growth and the yeast will just ferment the starter."

And in fact this agrees with 'Optimising Yeast Starters' as linked by @RoomWithABrew - which is a really good read by the way

 

[The-Engineer-That-Brews]

He goes on to say:

1:10 dilution is what I find to be the best dilution to obtain the maximal yield in yeast growth. This number can vary from person to person. Some say 1:5 others 1:20. The 1:10 dilution gives you the ideal cells/ml once a starter has reached confluency. I will bet serious money that this is how WYeast and white labs prep there yeast.

Unfortunately, adding more yeast, as in a 1:5 dilution, will not grow more yeast. At least this is what I see when I make my starters. Yeast will only grow (replicate) to the point where there is still sugar and nutrients in the medium. Add more yeast (1:5 dilution) just depletes those reserves faster.

If you want to make the starter with less steps, the only way to do it, efficiently, is to make a bigger sized starter with more yeast. 2 smack packs in a 3 liter starter for example.

 

[DocAnna]

Yeast will only grow (replicate) to the point where there is still sugar and nutrients in the medium. Add more yeast (1:5 dilution) just depletes those reserves faster.

The cells will still grow and divide if oxygen is available once the nutrients are depleted but will portion out their existing nutrients with each division until no longer viable or dormant. The exception oddly being when the glucose concentration is high at the beginning or with a stronger wort when the Crabtree effect applies and the cells don't divide but produce alcohol even in the presence of enough nutrition, bizarre but happens when there's lots of nutrition around and the cell can more quickly produce ATP (the energy currency) using the anaerobic pathway even though it's less efficient that the oxidative pathway. It's one of the reasons TETB's directions include not letting the starter go too far, as while there might be more yeast cells in the flask, they risk being less viable and increased incidence of petite mutants which are depleted of mitochondrial DNA and can't ferment properly.

Anyway... why not innoculate a tiny number of cells into a huge wort: since no matter how brilliant your sterile technique is, there's always going to be a small number of similar wee beasties that want to race your desired yeast cells to conquer the wort. Gradual step up allows domination by your yeast, and checking of it before the next step. There's a good slide presentation here on different ratio's of step up at different stages: https://www.vlb-berlin.org/sites/default/files/2018-11/04_Marshall_YeastManagement.pdf

 

[The-Engineer-That-Brews]

There's a good slide presentation here on different ratio's of step up at different stages: https://www.vlb-berlin.org/sites/default/files/2018-11/04_Marshall_YeastManagement.pdf

Interesting how they suggest using brewing wort for propagation as the hop content will deter bacterial growth... I suppose one could achieve the same effect by boiling some hops in the flask along with the DME...?