Anyone know how to step up frozen yeast and make a starter?

[rtstrider]

The krausen has dropped on the 1.5L 1.020 starter. In lieu of decanting I'm just going to add a half liter of wort to bring this up to 2L 1.040 this evening. I'm using the brewers friend yeast starter calculator. Per the calculator to make 2L of 1.040 wort you would need 2L of water and 213 grams DME. I'm going to add 213 grams DME to .5L water and boil for at least 5 minutes. Depending on how it looks I may go longer. I just don't want to have .5L of burnt DME syrup to deal with lol This Sierra Nevada Chico is stepping up significantly faster than it did from bottle dregs!

 

[IslandLizard]

The krausen has dropped on the 1.5L 1.020 starter.

Are you using a stir plate?

 

[DuncB]

Trying to see if I have any viability from a 14 month out of date WLP 1983 yeast.
Started with 800ml of 1.020 at 19.5 celsius on a stirplate. My 3 litre flask didn't look like it would cope with only a couple of hundred ml.
Not 24 hours yet we'll see how it goes fingers crossed.
I suspect less viable yeast in here than from a frozen slant.
Does anyone add hops to their starter?

 

[IslandLizard]

Does anyone add hops to their starter?

I've been contemplating that. Mostly to keep lactobacillus and possible other infections away.
Boil the water with a few pellets of Warrior or so for 15-20', to get around 15-20 IBU, then add the DME.

Out of 4, same source, I had one starter going sour on me (with a thick pellicle) when kept in the fridge for several weeks, for an unintended extended cold crash . The other 3 were A-OK.

Then with another 2 starters (different yeast), I had one going flaky, but the source of that was already suspect. The other (different source) was fine.

 

[rtstrider]

Are you using a stir plate?

Yes! I boiled .5L of the crazy strong wort for 10 minutes. Just topped off the flask. Here we go! 2 liters 1.040 brew day is next Thursday so it should be fine to sit in the kegerator after this. Will sure help with decanting

 

[IslandLizard]

Yes! I boiled .5L of the crazy strong wort for 10 minutes. Just topped off the flask. Here we go! 2 liters 1.040 brew day is next Thursday so it should be fine to sit in the kegerator after this. Will sure help with decanting

That looks very promising!

 

[DuncB]

Mine is a rather grey colour and lacks any vibrancy at the moment, will update on the life support.
I have flipped between adding hops and no hops. Currently no hops in the starter attempt but could add with a later
wort addition.

 

[IslandLizard]

Mine is a rather grey colour and lacks any vibrancy at the moment, will update on the life support.
I have flipped between adding hops and no hops. Currently no hops in the starter attempt but could add with a later
wort addition.

Mine tend to start dull brown too.
I always wait for the color to change to significantly lighter, as in @rtstrider's picture, above. Then give it another day, or 2 (sometimes longer). The change in color could take anywhere from 2-7 days or even longer depending on how old (less-viable) the yeast is.

 

[DuncB]

@IslandLizard

I'll watch out for some tell tale signs, certainly no visible bubbles when I took this photo but perhaps a little less grey than I thought.

 

[homebeerbrewer]

Does anyone add hops to their starter?

I've been contemplating that. Mostly to keep lactobacillus and possible other infections away.
Boil the water with a few pellets of Warrior or so for 15-20', to get around 15-20 IBU, then add the DME.

Interesting idea. I recently bought a 21 qt pressure cooker to do canning, and one plan I have for it is doing all grain starters and canning them. I hadn't thought about adding hops, but that may be a good experiment to try. The pressure cooker will also be my "poor man's" autoclave for my yeast bank equipment.

Another phrase to ad to my list: Gardening, the most expensive way to get cheap vegetables. The pressure cooker was purchased to can my veggies from the garden, but it's gonna be dual triple use now.

 

[DuncB]

I often find that the runnings after the sparge volume is done can be over 1.020 and a litre or three ( yes agreed that's an issue I haven't solved yet ).
I've put this in a sanitised pet bottle in the freezer and then defrost boil and have a weak starter or something to top up for my starter.

 

[rtstrider]

Around an hour and a half later holy moly!

 

[rtstrider]

According to the brewers friend yeast starter calculator this was in the 120 billion cell range before the .5L addition. The calculator says I should be in the 420 billion range after this ferments out In all reality it’s all guess work unless you know how to really count yeast so who knows

 

[rtstrider]

@IslandLizard

I'll watch out for some tell tale signs, certainly no visible bubbles when I took this photo but perhaps a little less grey than I thought.
View attachment 735535

Id actually give it up to 7 days. I would look more for color than krausen. I think you may have overdone it with the starter wort size though. Only thing to do now is hurry up and wait. Good luck!

Side note, how long can frozen liquid yeast last? I’ve found a ton of articles on how to make a liquid yeast bank but I have not found a time frame. I know it’s recommended to not let refrigerated slurry hang out for more than 6 months. So for frozen is it a year or two or is it many years?

 

[DuncB]

According to the brewers friend yeast starter calculator this was in the 120 billion cell range before the .5L addition. The calculator says I should be in the 420 billion range after this ferments out In all reality it’s all guess work unless you know how to really count yeast so who knows

Haven't you bought a microscope? I thought they were de rigeur for the yeast farmer and the way to work out the number of yeast cells.

 

[Brooothru]

The krausen has dropped on the 1.5L 1.020 starter. In lieu of decanting I'm just going to add a half liter of wort to bring this up to 2L 1.040 this evening. I'm using the brewers friend yeast starter calculator. Per the calculator to make 2L of 1.040 wort you would need 2L of water and 213 grams DME. I'm going to add 213 grams DME to .5L water and boil for at least 5 minutes. Depending on how it looks I may go longer. I just don't want to have .5L of burnt DME syrup to deal with lol This Sierra Nevada Chico is stepping up significantly faster than it did from bottle dregs!

That ought to work fine. I hope so, 'cause that's actually what I did on the batch I'm brewing right now: a Kolsch using WLP-029. Ran out of time so I pitched 1.5L 1.040 wort into the flask of 250 ml 1.020 last night. Good krausen this morning and brewing right now, so no time to cold crash and decant. There'll be ~ 1.75L of yeast slurry/supernatant going into the fermenter along with the wort. The yeast in the flask smelled wonderful this AM, so fingers crossed that ther won't be off-flavors in the fermenter.

 

[IslandLizard]

Side note, how long can frozen liquid yeast last?

Many, many years, as long as it doesn't defrost in between. Lab freezers go down to -20 to -40C or even lower.

Slants are the preferred way of long term storage. Our usual method using a glycerine mix in a (small) centrifuge tube is a makeshift way to prevent damaging ice crystals from forming, rupturing cells.

I know it’s recommended to not let refrigerated slurry hang out for more than 6 months.

Because viability goes down as usual, 10% per month or so. I've used slurries that have been refrigerated for 2 years and longer. The 2-3 year-old ones bounced right back in a 1.020 starter.

 

[timkeen]

I'm probably doing something "wrong", but I've been successfully using yeast from a frozen bank for a little over a year now. My process is essentially the glycerine/distilled water process described earlier in this thread, using 15 ml tubes kept in a non-frost-free deep freezer.

I live in south Florida where access to liquid yeast is problematic, and with 80+ degree ground water, so is chilling the wort post-boil. On brew day, I pull 1-2 vials of yeast from the freezer and let them thaw on the counter during the course of the brew. After boil is complete, I pump about 1200-1500 ml of still-boiling-hot wort into a 2L flask, chill it, and put it on a stir plate. Then I pitch the yeast (which has been thawing for several hours), making a real wort starter. The rest of the wort gets pumped into a hot cube and allowed to cool naturally overnight, or sometimes two if the starter needs more time. Once the starter is ready, they all join hands in a vessel in the ferm chamber.

I've had a few times where the starter never appeared to "come alive", but gravity tests and subsequent fermentations have so far worked out every time. The original bank samples were built using liquid packs and a DME starter, but every brew day uses one of the frozen children and a real wort starter, and I've never taken the time to step up the starters, either. I just pitch 1-2 vials (15-30ml) into roughly 1.5L wort and go from there. Someone is bound to excoriate me on underpitching/stressing/etc, but the results have been good, and a year later the frozen yeast is still viable.

For the record, I'm not using really exotic yeasts or making really big beers. I tend to make lower-gravity beers in the 1.04x-1.05x range: English porters, stouts, and browns, German hefes and kolsch, Belgian wits, and the occasional IPA or "lawnmower" beer. It's possible I might be unknowingly getting away with a bad practice.

 

[rtstrider]

That ought to work fine. I hope so, 'cause that's actually what I did on the batch I'm brewing right now: a Kolsch using WLP-029. Ran out of time so I pitched 1.5L 1.040 wort into the flask of 250 ml 1.020 last night. Good krausen this morning and brewing right now, so no time to cold crash and decant. There'll be ~ 1.75L of yeast slurry/supernatant going into the fermenter along with the wort. The yeast in the flask smelled wonderful this AM, so fingers crossed that ther won't be off-flavors in the fermenter.

Funny you mention wlp029. I pitched 2L of full starter Imperial G03 (supposedly the same strain) into 5.5 gallons of wort. Turned out way too good! Couldn’t tell. I just adjust the preboil wort size when I do that so I end up right where I want to be after adding the 2L. You’ll be just fine

 

[Brooothru]

Funny you mention wlp029. I pitched 2L of full starter Imperial G03 (supposedly the same strain) into 5.5 gallons of wort. Turned out way too good! Couldn’t tell. I just adjust the preboil wort size when I do that so I end up right where I want to be after adding the 2L. You’ll be just fine

Yeah, this is actually my first time using WLP-029. I'm not sure why, 'cause I really like a good Kolsch. Anyway, the yeast propagated quite nicely from a slightly older Pure Pitch package (it was slightly past its Best By date), but it responded quickly to a 1.020 vitality starter and kept coming on strong with the next 2 step ups. The 1.5L/1.040SG step really smelled nice, so I felt good about chucking the whole flask into the fermenter. Fast-forward 12 hours post pitch and she's really going to town at 62F. White Labs' spec sheet hints that its performance starts to suffer below 62F and can stall out early if you don't raise the temperature towards the end of fermentation. Got my FFT sample on the stir plate s we speak to get a prediction of how low it will go with this wort and this yeast batch, and my plan is to start slowly letting the temperature free-rise to 68~70F when I'm within 10 points of terminal gravity.

Unfortunately my unitank has a Helles that is spunding right now so my low pressure conical has yesterday's Kolsch fermenting in it. I may do a low pressure transfer to a sanitized and purged keg and let it spund while free-rising to cellar temperature (68-70F this time of year) when it's 10 points from being done, then cold crash the keg in the beer fridge to let it settle and condition. With a floating dip tube I hope to be able to then transfer most of it to a clean serving keg. Never fermented in a keg before, so this will be another first for me.

 

[rtstrider]

The krausen has not fallen yet on the 2L starter. It’s been 48 hours now. Guess it has a bit more to go

 

[IslandLizard]

The krausen has not fallen yet on the 2L starter. It’s been 48 hours now. Guess it has a bit more to go

Pitching an active starter has a lot of merit. Lift off within mere hours.

 

[DuncB]

I've added the second lot of wort to the wLP 1983 and it's fizzing away so it's come back to life.

 

[rtstrider]

Pitching an active starter has a lot of merit. Lift off within mere hours.

The krausen is finally hinting at falling. It’s been over 72 hours now. The starter on the stir plate is looking more like it should. More haze and less splotchy yeast/trub. The grains should be here tomorrow. Not brewing till next weekend though. Primary fermenter has an Anchor Liberty clone with wyeast 1272. Yeast starter was pitched around 9 days ago. Was worried something was up with that batch. When I pulled out the blow off hose to replace it with the 3 piece airlock the aroma was nice Did some reading and thinking 1272 is just a dog slow yeast. No biggie though. As long as I can get this kegged and brew by next Sunday I’m good Have nothing on my hands but time! Good problem to have lol

 

[rtstrider]

Yeah, this is actually my first time using WLP-029. I'm not sure why, 'cause I really like a good Kolsch. Anyway, the yeast propagated quite nicely from a slightly older Pure Pitch package (it was slightly past its Best By date), but it responded quickly to a 1.020 vitality starter and kept coming on strong with the next 2 step ups. The 1.5L/1.040SG step really smelled nice, so I felt good about chucking the whole flask into the fermenter. Fast-forward 12 hours post pitch and she's really going to town at 62F. White Labs' spec sheet hints that its performance starts to suffer below 62F and can stall out early if you don't raise the temperature towards the end of fermentation. Got my FFT sample on the stir plate s we speak to get a prediction of how low it will go with this wort and this yeast batch, and my plan is to start slowly letting the temperature free-rise to 68~70F when I'm within 10 points of terminal gravity.

Unfortunately my unitank has a Helles that is spunding right now so my low pressure conical has yesterday's Kolsch fermenting in it. I may do a low pressure transfer to a sanitized and purged keg and let it spund while free-rising to cellar temperature (68-70F this time of year) when it's 10 points from being done, then cold crash the keg in the beer fridge to let it settle and condition. With a floating dip tube I hope to be able to then transfer most of it to a clean serving keg. Never fermented in a keg before, so this will be another first for me.

One thing I noticed about G03 is the yeast is very tart while still in suspension. It clears nicely though. I fermented around 60F and was just fine Just giving a heads up in case you pick up the same tartness. It will absolutely disappear with gelatin fining and cold conditioning

 

[DuncB]

Third feeding after decanting a litre and a half and topping up activity complete looks like there was viable cells in that 16month post use date WLP 1983.

Think I've got plenty of cells here for my brew.

 

[rtstrider]

Well the 2L starter finally fermented out. I'm not putting too much weight into the propagation time on this batch because the Sierra Nevada Pale Ale Yeast (@Northern_Brewer ) usually is dog slow from my experiences. It's clean though and ready to go for brew day. I just put the flask in the fridge. The only thing that is going to hold back brew day is the fermenter with 1272 in there. It's finishing up very slowly. I fermented at 63F and as of Saturday morning I've been raising the temp 1F per day until it hits 68F. This is one of those times I wish I had a second fermenter...I do have some old brew buckets laying around lol

 

[Northern_Brewer]

the Sierra Nevada Pale Ale Yeast (@Northern_Brewer )

 

[InspectorJon]

Here might be a good place to ask this question. As I understand it fermentation kind of goes in two phases. Cell growth then fermenting sugar into alcohol. I'm guessing there's some of each going on at the same time but out goal is healthy yeast, not alcohol when building up starters. Do we really want our starter wort to ferment out all the way or is there a more optimal time to cold crash the yeast out and move to the next step?

 

[rtstrider]

Got the slurry pitched into a nice old school IPA about 2 hours ago now. I absolutely love the Sierra Nevada Pale Ale Yeast when it comes to pale ales and IPAs! Going to be getting a w34/70 tube started in the morning. An Oktoberfest is the next brew I’ll keep everyone posted on the status of the frozen Sierra Nevada yeast brew!

 

[rtstrider]

Well so far I’d call this a success! Just over 24 hours and we have krausen!

 

[DJTroz]

I hate to revive a 6 month old thread but wish I had found this yesterday. I have been building up a bank of about 9 different strains using the Iso alchohol freezing method glycerine storage solution using ~5ml slurry to 10ml glycerine solution. I have finally decided to start using one of them. I searched for a few days trying to find the method to build from a frozen sample but out of everything I searched, I couldn't find anything. The only article I found was from a different source where it indicated to thaw to around 80 degrees as quickly as possible then pitch like normal (1.036-1.04, 1l). I knew this sounded odd for two reasons; one to defrost this quickly didn't seem right (but I am an analyst, not biologist, so not sure), and the other that when you work with a small sample, you start with a lower volume of wort and lower gravity. In either case, knowing I had another 10 vials of this strain, I took a chance and used 3 tubes (15ml of slurry with the 30ml of glycerine solution), dropped in a container of water to bring to around 70 degrees, then pitched in 1l of 1.036. This was around 6p yesterday. This morning, it had a good krausen, smells normal, typical creamy color, and have a small amount of yeast settling once I took it off of the stirplate.

If I'm reading this thread correctly; you guys are doing

Slowly bring tubes from frozen state to room temp over 1-4 days (each person's method slightly different). When pitching frozen culture, pitch all of contents (slurry and glycerine).

Stepping Method:

1. 750-1000ml of 1.02, then either crash/decant or simply add to step 2
2. Step to 1.5l of 1.02, then crash/decant
3. For the third step, treat like a normal starter as if it was a fresh pack of yeast, or did I miss something somewhere?

Does anyone feel that there is more value in a longer warming period (days) versus hours? I would think it is less stress on the cells, but again, I don't know either.

I'm probably going to start another iteration using the new method, as well as keep the original starter going to compare the differences.

 

[rtstrider]

I hate to revive a 6 month old thread but wish I had found this yesterday. I have been building up a bank of about 9 different strains using the Iso alchohol freezing method glycerine storage solution using ~5ml slurry to 10ml glycerine solution. I have finally decided to start using one of them. I searched for a few days trying to find the method to build from a frozen sample but out of everything I searched, I couldn't find anything. The only article I found was from a different source where it indicated to thaw to around 80 degrees as quickly as possible then pitch like normal (1.036-1.04, 1l). I knew this sounded odd for two reasons; one to defrost this quickly didn't seem right (but I am an analyst, not biologist, so not sure), and the other that when you work with a small sample, you start with a lower volume of wort and lower gravity. In either case, knowing I had another 10 vials of this strain, I took a chance and used 3 tubes (15ml of slurry with the 30ml of glycerine solution), dropped in a container of water to bring to around 70 degrees, then pitched in 1l of 1.036. This was around 6p yesterday. This morning, it had a good krausen, smells normal, typical creamy color, and have a small amount of yeast settling once I took it off of the stirplate.

If I'm reading this thread correctly; you guys are doing

Slowly bring tubes from frozen state to room temp over 1-4 days (each person's method slightly different). When pitching frozen culture, pitch all of contents (slurry and glycerine).

Stepping Method:

1. 750-1000ml of 1.02, then either crash/decant or simply add to step 2
2. Step to 1.5l of 1.02, then crash/decant
3. For the third step, treat like a normal starter as if it was a fresh pack of yeast, or did I miss something somewhere?

Does anyone feel that there is more value in a longer warming period (days) versus hours? I would think it is less stress on the cells, but again, I don't know either.

I'm probably going to start another iteration using the new method, as well as keep the original starter going to compare the differences.

I just run luke warm water over the frozen tube until it thaws. I'll shake the tube and get it to room temp as soon as the first batch of starter wort is at pitching temps.

For the steps its

1. Pour 1 15ml tube (well shaken) into 150ml 1.020 wort with a pinch of yeast nutrient. This is 8 grams dme, pinch of yeast nutrient, and a quarter of crushed Campden tablet. The Campden is to convert/cook off any chlorine/chloramines. I use a sanitized pyrex measuring cup and bring it to a quick boil in the microwave. From there it's covered with foil and goes into the freezer for a half hour to hour on a pot holder. Usually that gets to pitching temps. This gets poured into a sanitized pint jar with the quick thawed slurry. I cover the pint jar with sanitized foil and wait anxiously for 72 hours or so for the first signs of activity. I'll just swirl the yeast into suspension each time I walk by. That's maybe 2 or 3 times a day tops. I don't shake on this step. This step is the longest. That goes until the krausen disappears. This nets in the 25-27 billion range worth of yeast cells

2. Get 1.5L 1.020 wort boiled with yeast nutrient, and another quarter Campden tablet, chill to pitching temps, then pour the whole 150ml jar into the 1.5L 1.020 wort and let er rip for 48-72 hours (sometimes it's shorter than that) on the stir plate until the krausen drops/disappears. The amount of DME required for this step is 79.9 grams or 2.8oz This ends up getting in the 100-115 billion cell range for the yeast cells (aka Wyeast smack pack)

3. This is the final step of 2L 1.040 wort plus yeast nutrient and another quarter Campden tablet. I just add 213 grams dme to 500ml water plus pinch of yeast nutrient and boil that for 15 min, chill to pitching temps, then top off the 2L beaker on the stir plate. Again this goes until the krausen disappears. From there it gets an overnight cold crash and decant/poured into whatever's brewed the next day. This supposedly jumps to the 400 billion cell range (aka 2 Imperial yeast packets or 4 Wyeast smack packs)

In total this takes around 1.5 weeks to 2 weeks from the initial quick thaw to get to a pitchable count. According to brewersfriend this nets around 400 billion cells. In reality it's a crapshoot unless you're willing to count the cells. I'll also say this much. My brews have been significantly better after fermentation since stepping up the freezer vials. Hopefully this helps!

 

[Brooothru]

I just run luke warm water over the frozen tube until it thaws. I'll shake the tube and get it to room temp as soon as the first batch of starter wort is at pitching temps.

For the steps its

1. Pour 1 15ml tube (well shaken) into 150ml 1.020 wort with a pinch of yeast nutrient. This is 8 grams dme, pinch of yeast nutrient, and a quarter of crushed Campden tablet. The Campden is to convert/cook off any chlorine/chloramines. I use a sanitized pyrex measuring cup and bring it to a quick boil in the microwave. From there it's covered with foil and goes into the freezer for a half hour to hour on a pot holder. Usually that gets to pitching temps. This gets poured into a sanitized pint jar with the quick thawed slurry. I cover the pint jar with sanitized foil and wait anxiously for 72 hours or so for the first signs of activity. I'll just swirl the yeast into suspension each time I walk by. That's maybe 2 or 3 times a day tops. I don't shake on this step. This step is the longest. That goes until the krausen disappears. This nets in the 25-27 billion range worth of yeast cells

2. Get 1.5L 1.020 wort boiled with yeast nutrient, and another quarter Campden tablet, chill to pitching temps, then pour the whole 150ml jar into the 1.5L 1.020 wort and let er rip for 48-72 hours (sometimes it's shorter than that) on the stir plate until the krausen drops/disappears. The amount of DME required for this step is 79.9 grams or 2.8oz This ends up getting in the 100-115 billion cell range for the yeast cells (aka Wyeast smack pack)

3. This is the final step of 2L 1.040 wort plus yeast nutrient and another quarter Campden tablet. I just add 213 grams dme to 500ml water plus pinch of yeast nutrient and boil that for 15 min, chill to pitching temps, then top off the 2L beaker on the stir plate. Again this goes until the krausen disappears. From there it gets an overnight cold crash and decant/poured into whatever's brewed the next day. This supposedly jumps to the 400 billion cell range (aka 2 Imperial yeast packets or 4 Wyeast smack packs)

In total this takes around 1.5 weeks to 2 weeks from the initial quick thaw to get to a pitchable count. According to brewersfriend this nets around 400 billion cells. In reality it's a crapshoot unless you're willing to count the cells. I'll also say this much. My brews have been significantly better after fermentation since stepping up the freezer vials. Hopefully this helps!

Agree with everything ^^above^^ with the possible exception of thawing. I prefer to let the sample sit in the refrigerator for a day or two to thaw slowly. The yeast have already been stressed enough, it seems to me. I don't have any 'scientific' reason or proof for preferring this methodology, other than "that's the way I do it when I'm thawing thick steaks or a prime rib." Force of habit and superstition I guess.

 

[rtstrider]

Agree with everything ^^above^^ with the possible exception of thawing. I prefer to let the sample sit in the refrigerator for a day or two to thaw slowly. The yeast have already been stressed enough, it seems to me. I don't have any 'scientific' reason or proof for preferring this methodology, other than "that's the way I do it when I'm thawing thick steaks or a prime rib." Force of habit and superstition I guess.

I haven't noticed any negatives to quick thawing at all. If anything it means the yeast is pitched a day or two sooner Btw I was reading through this thread and saw your mention of WLP029. Coincidentally guess what's on the final leg of being stepped up on the stir plate as I type....Imperial G03 lol That was actually cultured up from my homebrew bottles last March as a test. The batch was fined with gelatin and everything. This yeast has come back and performed exceptionally well on the starter steps! Hoping to rebrew the "Omega Lutra Kolsch" tomorrow except use the Imperial G03 yeast

 

[Brooothru]

I haven't noticed any negatives to quick thawing at all. If anything it means the yeast is pitched a day or two sooner Btw I was reading through this thread and saw your mention of WLP029. Coincidentally guess what's on the final leg of being stepped up on the stir plate as I type....Imperial G03 lol That was actually cultured up from my homebrew bottles last March as a test. The batch was fined with gelatin and everything. This yeast has come back and performed exceptionally well on the starter steps! Hoping to rebrew the "Omega Lutra Kolsch" tomorrow except use the Imperial G03 yeast

Might as well. It already has the fining gelatin in it!