# From Slant To Starter - In Pictures



## Wolfy (16/7/10)

As with most things yeast related, there are many ways to achieve the same result, but here the process I use to reculture yeast from a slant and grow it up to a starter of suitable pitching volume.
In this example I'm using a 6m old home-made slant of Wyeast Klsch yeast (2565) and I wanted to make a starter that was 2L in volume.
The starter was stepped up from about 15ml, to 100ml, 400ml and then 2L.

First step was to let the slant come up to room temp, gather the ingredients and clean and clean/sanitise everything:






The saved wort I had kept in the fridge for a couple of months was a bit gunky, so I made the starter wort by boiling about 175g of LDME (and some yeast nutrient) in a little more than 2L of water - aiming for an OG a bit above 1.030.
While the wort was still boiling hot I carefully tipped it into the various containers, topped them with aluminum foil and left them to cool:





The 15ml test tube of wort was quickly cooled in some cold water and then added to the slanted yeast.
Using a (plastic) inoculation loop I carefully scraped the yeast from the surface of the slant, trying to remove all the yeast but leave the solid agar mix behind:





After tipping the 15ml wort and yeast back into the clean test tube, I kept it on my computer desk, so I'd be reminded to shake it often.
The temperate was generally about 20C and the stater was allowed to ferment out for 24 hours, before stepping up to 100ml:




There is already a small layer of yeast on the bottom of the test tube.
At this stage due to the very small volume of beer a smell/taste test is not going to tell you much, as I suspect there were a few dead yeast cells scraped off the slant.

The 100ml starter was put onto my el-cheapo DIY stir plate and allowed to ferment out for another 24h, before stepping it up to 400ml:




The foam on the yeasty-starter on the left is from yeast fermentation, and the plain wort on the right from me shaking the crap out of it for about 15 mins before pitching the yeast.
(I had a little left over wort so I added some agar agar and made the new blank slants that can be seen in the background.)

After another 24h on the stir plate, the 400ml starter had created a nice layer of yeast, which I left to settle out for a little while so I could take a taste/smell sample:




Klsch yeast is not the nicest tasting stuff so I wanted to try to sample some beer without too much yeast in it. 

After it had passed the taste-test it was pitched into the final 2L volume:





The yeast was still crawling out of the top of the flask after 24 hours, so I left it on the stir plate for another day to let it complete fermenting.




The 9l square plastic container may look ugly, but there is no way the yeast can over-flow it and short out the electronics in the stir plate. 

After one more day - this time in the fridge:





Any yeast that did not escape in the previous step settled to the bottom of the flask:




The spent beer is now ready to be decanted, so yeast can be pitched into a batch of beer.


----------



## Pete2501 (16/7/10)

Nice post Wolfy. :beerbang:


----------



## ~MikE (16/7/10)

nice work, IMO considering you're using a stirplate you could happily go slant -> 50ml culture -> 1500ml starter.


----------



## Wolfy (16/7/10)

~MikE said:


> nice work, IMO considering you're using a stirplate you could happily go slant -> 50ml culture -> 1500ml starter.


Most likely true, but if I'm not in a hurry I do tend to be a little conservative, and play things on the safe side.
I was also slightly concerned about the yeast/slant viability, especially when the first step (and the second to a lesser extent) had a noticeable burnt-rubber smell (I suspect from autolysis of the yeast on the slant) .


----------



## bjay (16/7/10)

This is going straight to the pool room 
Thanks wolfy very infomative

cheers
bjay


----------



## WarmBeer (16/7/10)

Wolfy,

The wort in the 2lt flask looks considerably darker than the other 2.

Is this just due to the depth of the wort, or a different batch?

PS. Awesome guide. :kooi:


----------



## MeLoveBeer (16/7/10)

Thanks Wolfy, we're now starting to build up a great collection of yeast related threads; there is no excuse now for those wanting to split, farm, wash and start yeast.


----------



## NickB (16/7/10)

Perfect timing - I'm going to be culturing up a starter from a slant I was given sometime today, and was really trying to get my head around the process. Thanks Wolfy!

Cheers


----------



## praxis178 (16/7/10)

Funny I finished a sequence just like that yesterday (pitched 4L starter for two 23L ales) and my Mum stopped by as I was washing the cake from my Irish Red (WY 1084) afterwards, she watched me for a few mins and then said that her old boss at the yeast factory might be wanting to get in contact with me later. Couldn't have asked for a better compliment.....


----------



## geoff_tewierik (16/7/10)

Congrats on taking the time and putting in the effort to put the pictyure show together.

I did cringe at the addition of wort to the slant and scraping the yeast off, it's a waste of a good slant IMO.

Preferred method would have to be to take, asceptically, a small amount of yeast from the slant with an innoculating loop and add that to your first bit of wort. This will allow you to retain the slant for future brews/comparison tests and allow you to maintain a pure culture strain for a longer period of time.

The above comment is based entirely on there being just the one slant, if you've got a bunch of the same yeast on a shed load of slants then it's a non issue for you.

YMMV.

Cheers,

GT


----------



## Fourstar (16/7/10)

Beautiful work wolfy. Great commitment to document w/photos all of that over a few days. More effort than what i would put in! Pretty much the exact same procedure i perform except all of my erlenmyers have been smashed. Two of them from vibrations from the screws mounting my fan on the DIY stirplate. These have now been taped over to save any future flasks. <_< The past two ive been doing in some 1L and 2L fowles varcola preserving jars and are doing the same job. only issue is boiling the starter wort in a little saucepan.

The difference i have in my procedure too is i will just dump the 30ml or so of wort into the testtube, shake and leave to ferment out, once done i shake again and transfer into my flask. unless its one of the early crappy gelatine slants i made, it seems to work well without taking all the agar with it. :lol:


----------



## QldKev (16/7/10)

Good write up. I think it should get a few more people interested in farming yeast. I've always just used test tubes of yeast. What would be the pros/cons of slants over a test tube of yeast?

QldKev


----------



## Sammus (16/7/10)

~MikE said:


> nice work, IMO considering you're using a stirplate you could happily go slant -> 50ml culture -> 1500ml starter.



you probably could, but they say the max step for optimal growth should be between 5x and 10x. I do slant -> 10ml - 50ml - 250ml - 2.5L for lagers, or 10ml - 100ml - 1L for ales.


----------



## ~MikE (16/7/10)

Sammus said:


> ...but they say...


who says? h34r: 

you're probably right though, but given he's using a stirplate, it means he could easily afford to be lazy - if your starter vessel can maintain optimal growth conditions with agitation and added oxygen, and you're satisfied with your sterilisation procedures you could happily step up to up to ~ 50X i think.


----------



## WarmBeer (16/7/10)

Fourstar said:


> The difference i have in my procedure too is i will just dump the 30ml or so of wort into the testtube, shake and leave to ferment out, once done i shake again and transfer into my flask. unless its one of the early crappy gelatine slants i made, it seems to work well without taking all the agar with it. :lol:


So the wort in the test-tube doesn't disolve the agar overnight?


----------



## Fourstar (16/7/10)

WarmBeer said:


> So the wort in the test-tube doesn't disolve the agar overnight?



nope. its melting point is like 95 deg from memory.


----------



## cubbie (16/7/10)

So how many yeast cells do you estimate you have in the final starter?


----------



## Sammus (16/7/10)

~MikE said:


> who says? h34r:
> 
> you're probably right though, but given he's using a stirplate, it means he could easily afford to be lazy - if your starter vessel can maintain optimal growth conditions with agitation and added oxygen, and you're satisfied with your sterilisation procedures you could happily step up to up to ~ 50X i think.



I knew someone would say that 

I've been searching for the article since I posted that...there was some research that sounded convincing that concluded what I said... now just to find it


----------



## WarmBeer (16/7/10)

Fourstar said:


> nope. its melting point is like 95 deg from memory.


Ah, noice.

Hopefully you and Wolfy can give some hints and tips on the subject at the swap tomorrow.

Beginning to seriously wish I'd paid more attention during high school Chem prac.


----------



## ~MikE (16/7/10)

Sammus said:


> I knew someone would say that
> 
> I've been searching for the article since I posted that...there was some research that sounded convincing that concluded what I said... now just to find it



hehe, no no i believe you. in theory stepping it up on a smaller scale would definitely the safest way to go. in practice the difference would be negligible with optimum growth conditions and good sterilisation.



Fourstar said:


> nope. its melting point is like 95 deg from memory.


yups, it melts at ~85C (although i take it to near boiling point to get it to dissolve properly) and solidifies at ~40C and it's a disaccharide polymer that isn't metabolised, making it far better than gelatine which will liquify in the presence of proteases.


----------



## Wolfy (16/7/10)

WarmBeer said:


> The wort in the 2lt flask looks considerably darker than the other 2.
> 
> Is this just due to the depth of the wort, or a different batch?


Yep it's just the depth of the wort and probably related to the camera flash etc, all the flasks were filled with the same wort.


geoff_tewierik said:


> I did cringe at the addition of wort to the slant and scraping the yeast off, it's a waste of a good slant IMO.
> 
> Preferred method would have to be to take, asceptically, a small amount of yeast from the slant with an innoculating loop and add that to your first bit of wort. This will allow you to retain the slant for future brews/comparison tests and allow you to maintain a pure culture strain for a longer period of time.
> 
> The above comment is based entirely on there being just the one slant, if you've got a bunch of the same yeast on a shed load of slants then it's a non issue for you.


Yes, if I only had one slant I'd be much more careful, use the procedure you mentioned and probably a smaller amount of wort for the first step. However I generally make 3 or 4 slants of each yeast I have, this seems to work well for me in that I usually have 1 left after about a year when it's time to reslant that strain.
I'll be following your procedure with the (one) commercial slant I have here, someone has said that they have been reusing the same slant - just taking a small amount of yeast each time - for 4 years now. 


QldKev said:


> I've always just used test tubes of yeast. What would be the pros/cons of slants over a test tube of yeast?


Test tubes of yeast would require less preparation, but I think the yeast stored on wort/agar is a little more stable and once made they are easy to handle and store.
You can also create a slant from a small sample of 'virgin' generation yeast - from a streak plate for example - and it's easy to reslant old samples so that the yeast always remains viable, with test tube storage you'd need to grow and split the yeast each time you needed to reculture it.
Splitting new packs of Wyeast into test tubes via *Tony's* method (or similar) is much easier (and maybe better) than using slants, but it really 'only' lets you split the pack into a limited number of test tubes, once those are used you'll need to buy a new pack.
With slants (and streak plates) you should be able to 'save' that particular strain of yeast for an indefinate period.


cubbie said:


> So how many yeast cells do you estimate you have in the final starter?


Personally, I tend not to over-complicate things in this regard, so I make some big assumptions in regard to the cell count.
I assume that since I am using small steps, a stir plate and a healthy dose of yeast nutrient, at each stage I'll get maximum growth from the yeast.
Then I let someone Else's maths do the work for me, and see what size starter is recommended by the MrMalty calculator (assuming 1 pack of yeast and maximum growth).
Assuming again that the second-to-last-step of the starter process should have about the same number of yeast cells as commercial pack/vial of yeast (_I don't recall where the basis for that theory came from_), I base my starter volume on what the calculator suggests - usually rounded up to the next convenient size container I have.


----------



## Fourstar (16/7/10)

Sammus said:


> I knew someone would say that
> I've been searching for the article since I posted that...there was some research that sounded convincing that concluded what I said... now just to find it



I think thats research Jamil Z and Whitelabs have undertaken. I remember hearing that.


----------



## Wolfy (16/7/10)

~MikE said:


> yups, it melts at ~85C (although i take it to near boiling point to get it to dissolve properly) and solidifies at ~40C and it's a disaccharide polymer that isn't metabolised, making it far better than gelatine which will liquify in the presence of proteases.


Agar agar is great, when making the slants shown in the background of the photos, I usually heat it in the microwave for 30 seconds then stir to dissolve. However, I was a little impatient and shoved it in for 1min and then went to the bathroom, when I came back it had created a huge sticky boiling mess by overflowing the jug. By the time I had finished making the slants (fun stuff first clean up later) the overflowing agar mess in the microwave had solidified and the only clean up required was to literally peel off the solid wort-jelly mess in one big sheet.
The same was true for cleaning up the saucepans, jugs and syringe, while I did shove them in a sink full of warm soapy water and let them soak, the agar-wort remained as a solid mass that was easily peeled off each item.

Note: After the I add the agar-wort mixture to the test-tubes I give them a decent amount of time in the 'pressure cooker' so mixing the agar in the microwave does not really compromise the (close to) sterilization procedures required when making slants. The Geelong/Hawthorn football match was exciting to watch, so I only remembered the slants were cooking when I went back into the kitchen after the match had finished.


----------



## wakkatoo (17/8/10)

subscribed


----------



## haysie (17/8/10)

Wolfy, super threads, after all those slants you made....... what are ya gotta do with them? 
You cant make that much beer in a year let alone drink it. The very reason slanting aint for me, I like to split, use and move on.


----------



## Wolfy (17/8/10)

haysie said:


> Wolfy, super threads, after all those slants you made....... what are ya gotta do with them?
> You cant make that much beer in a year let alone drink it. The very reason slanting aint for me, I like to split, use and move on.


You're right, it is getting a bit that way.
But I do use about 5 slants for each new strain (duplicates for the initial culture, then 3 more once I isolate a sample, 2 of those go in the fridge and 1 is used up to make a frozen sample) so they actually get used quickly.
It's also nice to have specific yeast on hand if/when I want it - you're welcome to visit if you ever need Pacman or 1469 for example. 
Slants also allow for the 'banking' of yeast that is not (easily, often or usually) commercially available (I hope to have some of the unique old-Brewtek European yeast soon, for example).

However, other than that, for many/most people splitting new yeast packs/vials into sterile tubes (as you do) works well, is easier and much less maintenance.


----------



## dcx3 (20/8/10)

Just curious if any yeast gurus have ever tried to harvest the yeast in coopers vintage ale.


----------



## wakkatoo (10/9/10)

Just ordered some tubes, loops and flasks from ProSciTech. Only other thing now is a 2l Flask as they didn't have them and to source some agar, which I'm assuming is not a hard thing?


----------



## manticle (10/9/10)

I think chinese grocers/food shops sell agar-agar


----------



## Golani51 (24/3/11)

Wolfy said:


> Yep it's just the depth of the wort and probably related to the camera flash etc, all the flasks were filled with the same wort.
> 
> Yes, if I only had one slant I'd be much more careful, use the procedure you mentioned and probably a smaller amount of wort for the first step. However I generally make 3 or 4 slants of each yeast I have, this seems to work well for me in that I usually have 1 left after about a year when it's time to reslant that strain.
> I'll be following your procedure with the (one) commercial slant I have here, someone has said that they have been reusing the same slant - just taking a small amount of yeast each time - for 4 years now.
> ...




What is Tony's method?


----------



## Wolfy (24/3/11)

Golani51 said:


> What is Tony's method?


The search function is your friend. 
http://www.aussiehomebrewer.com/forum/inde...showtopic=45866


----------



## Golani51 (24/3/11)

Wolfy said:


> The search function is your friend.
> http://www.aussiehomebrewer.com/forum/inde...showtopic=45866




Thanks champ


----------



## Bizier (24/3/11)

I am concerned with concentrations of nutrients, seeing as I tried to rehydrate with some generic nutrient once (wasn't thinking properly) and damaged/killed the yeast.

Can I get some advice on how much nutrient to use, say per Litre?


----------



## Wolfy (24/3/11)

Bizier said:


> I am concerned with concentrations of nutrients, seeing as I tried to rehydrate with some generic nutrient once (wasn't thinking properly) and damaged/killed the yeast.
> 
> Can I get some advice on how much nutrient to use, say per Litre?


Are you asking about rehydrating dry yeast, how much 'yeast nutrient' to use in a starter, or if/how much you should use in the first steps of stepping up from a slant (or from a small sample of stressed yeast)?


----------



## Bizier (25/3/11)

My understanding was that you mixed nutrient into a common wort for all your stepping purposes. I would just like to know a good amount to use for this. I understand that there are specific nutrients developed for rehydrating dry yeast, and that the generic stuff is not suitable.


----------



## Wolfy (25/3/11)

Bizier said:


> My understanding was that you mixed nutrient into a common wort for all your stepping purposes. I would just like to know a good amount to use for this. I understand that there are specific nutrients developed for rehydrating dry yeast, and that the generic stuff is not suitable.


The usage level for the (commercial-type) Yeast Food I have is 40ppm (40g/10hl), which is about 0.1g for starter sizes.
Since I prepare all steps of the starter at the same time (and at the same gravity) I just dip a teaspoon in the Yeast Food and throw a tiny bit in.
Having said that - in theory - the first steps of the starter should be in lower gravity (1.020) wort (presumably with less nutrients) if the starting yeast is old, tired or stressed (like the sample was in the pictures at the start of this thread).

I don't have any of the nutrients recommended for rehydrating dry yeast, so I just do that in plain water.


----------



## Bizier (25/3/11)

Cheers Wolfy.


----------



## Jarthy (8/6/11)

hope this isn't a silly question, as I'm getting quite interested in doing some yeast farming, but does light strike effect starters?


----------



## felten (8/6/11)

Lightstrike only affects the hop compounds, so it won't affect your starter if you aren't adding hops.


----------



## Jarthy (8/6/11)

that makes sense. thanks felten.


----------



## Thefatdoghead (5/12/11)

So Wolfy if I counted right it took you 6 days to go from slant to pitching?


----------



## Wolfy (5/12/11)

Gav80 said:


> So Wolfy if I counted right it took you 6 days to go from slant to pitching?


Yeah, that sounds about right, 5 to 7 days is what I usually allow.


----------



## MaltyHops (22/2/12)

From another thread but more appropriate here...



Wolfy said:


> The longest I generally leave each step of the starter-process on the stir-plate is 48hours
> (usually it's just 24h), and as I suggested have never noticed a problem.
> However, I also let the yeast settle, decant the spent starter and pitch only the yeast, and
> use stepped-starters (which would minimise the presence of dead yeast) both of which may
> or may not be the same procedures that others use.


Gday Wolfy, 

What if the the yeast are still active after 48 (or 24) hours? Do you let them ferment out
after taking them off the stirplate or into the fridge to force them to settle?

From what I've come to understand, the yeast growth cell count is based on the amount of
food (SG & volume) in a starter so if growth is stopped before time, the expected cel count
wont be reached.

Actually, I seem to recall reading somewhere that the bulk of yeast reproduction happens
in the first 12 hours - is this what you're basing your method on?


----------



## Wolfy (22/2/12)

Yeast 'activity' is different to yeast growth, most yeast growth occurs within 12 hours, sometimes a bit longer longer for larger volumes.
However, 24h is a much more convenient time-period to step-up starters, and most if not all the yeast growth should be finished by that time, so that works for me.

Only at the last step of the starter-process do you want to consider letting the yeast ferment the starter fully, that way they build up their reserves making them more healthy when pitched into your beer-wort.


----------



## MaltyHops (22/2/12)

Trouble is there can often be still a fair bit of 'activity' after 48hrs 
(at least for the last few times I've been stir-plating) making it hard
to decant the beer off without losing quite a bit of yeast in suspension
(unless this doesn't really matter?).

Could stick the starter into the fridge to settle the yeast but would
the cool/warm/cool/... of a step up or two be bad for the yeast?


----------



## Wolfy (23/2/12)

When I say "_on the stir plate_" I mean on the stir plate with it turned on, spinning around, stirring and oxygenating the starter.

From one starter-step to the next I do not decant the spent wort, but pitch the entire starter into the next step up.

For larger size starters (greater than 2L) I might leave them on the stir plate for 2 days, however if it's the last step of the process it will be longer than that before it's ready to pitch.
On the last step only, I like to let it fully ferment and then give it time to settle out - that can only happen after the stir-plate is turned off, but how long it takes depends on the yeast and starter-size.

You could easily put that last starter step in the fridge to help it settle, but for me that depends on how quickly I'm going to use it.
If I've planned well and it will be used as soon as it's ready, I try not to put it in the fridge since that could cause thermal-shock to the yeast (if my fermenting fridge is set to the correct temp I'll put the starter in there to settle-out), but if I need to store it for a short time (hopefully a maximum of 1 week) then it goes in the fridge.


----------



## MaltyHops (23/2/12)

Ah yes, just realised I've muddied the waters because I'm using a single
1L stir-plate flask only so have to wait for the yeast to settle before
pouring off the starter beer before adding more wort. I guess I could
adapt my process to transfer 3/4 of each starter into a holding jar while
I add wort to the stir-plate flask to grow more yeast.

Thanks.


----------



## stux (23/2/12)

Consider investing in a family of erlenmeyers 

Everything except a 5L is fairly cheap anyway and its the shipping which kills you

Having a 500ml, 1L, 2L and 5L makes it sooooo easy to do steps!

Just pour one into the next and decant the final one

I like to keep my erlenmeyers less than 3/4 full


----------



## donburke (23/2/12)

wolfy, 

i notice you boil the full volume of the wort you will be using on day 1 and then splitting

some of this wort is not being used until then next day, and then again the day after, as you reach your next step

have you ever had a problem with the wort picking up any bugs waiting those couple of days to use it ?


----------



## Wolfy (23/2/12)

MaltyHops said:


> Ah yes, just realised I've muddied the waters because I'm using a single
> 1L stir-plate flask


You don't have to use flasks, anything of a suitable size with a flat-bottom works fine. I use demijohns for my large starters, it's just a bit difficult getting the stir-bar settled (and they are MUCH cheaper than 4-5 L flasks).


donburke said:


> have you ever had a problem with the wort picking up any bugs waiting those couple of days to use it ?


Never had a problem - so far - but I do not remove the alu-foil cap at all, just wrap cling-wrap around it, so the chance of external contamination is very small.


----------



## MaltyHops (23/2/12)

Wolfy said:


> donburke said:
> 
> 
> > have you ever had a problem with the wort picking up any bugs waiting those couple of days to use it ?
> ...


One time I boiled up a 1/2L starter wort, took about 10mL off (for a testtube slant)
while the wort was hot and put alfoil cap on the starter wort. The slant took a couple
of days to take off and I had left the capped wort on the kitchen bench the whole 
time which went off and had to be ditched. In hindsight, I think putting the starter
wort in the fridge would have prevented this.


----------



## Lyrebird_Cycles (11/7/16)

Error ignore this


----------



## Lyrebird_Cycles (11/7/16)

Error, ignore this


----------

