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I just use a mason jar and fill the alcohol to the level of the liquid in the tubes. The number of tubes and volume of alcohol don't seem to affect the viability noticeably. Some day I'll actually track the temperature drop in the tube to see how far off I am from the 1degC/min.

Even though DMSO would work as well, definitely not the right application here, and in case anyone else is reading this, DON'T USE DMSO! But as far as glycerin, I'm not too concerned. If you take that 5ml of 25% glycerin (1.25ml pure glycerin), and dump it all in the 1000ml starter, you would...

Any USP glycerin from the drug store will work. I would make a stock of the glycerin/water solution in a mason jar so you always have it on hand, or just put it into the tubes after it's cooled a bit, either works. You can do 30% glycerin or 25% glycerin and water (typo in the protocol), they...

I had that happen a couple of years ago. I refroze them, and only some of them made it back. One really screwed up a batch, which I attributed to some weird mutation, no bacteria, good starter, etc. I'd make starters and bring them back as quick as possible.

15ml

Yep, bacteria will do fine with all of these procedures. Nottingham will work fine as well. If you're banking to save some money on shipping, I would make a master batch right from the package and freeze. After a run through a stressful batch (most ciders/high gravity due to lacking nutrients)...

I use a small one that holds 4 quart size mason jars, so maybe it's 8qt? Plenty big for doing this kind of stuff.

It's all about standardizing your own protocol, and it looks like it works great for you! I've got some Conan in the freezer that I need to bring out for a brew, haven't had a chance to try it. I need to see what everyone is raving about...

Yeah, I don't think this guy is down with spreading the love. From an article on him and Fossil Fuels Brewing Co, "His only worry is that the unfiltered nature of this beer means that some of his yeast will invariably settle to the bottom of the glass or bottle, and an unscrupulous brewer could...

Any size will do, but get one that you can put a few mason jars in. And when I say pressure cook it, I mean in glass jars that you can then seal shut once the cook is done. Sorry if I made it seem like the cooker was full of solution, that's not what you want to do. I use a cheap aluminum one...

Agreed, we're not commercial, and it's very hard to standardize one rule of thumb for all strains, for all beers. Even so, I generally go with the standard commercial pitching rule of 0.75M/ml/deg plato for an ale and 1.5M/ml/deg plato for lagers (that's 10-20mil/ml for a 1.050 beer. That would...

1) Totally, you could definitely just freeze the smackpack or WL vial to a final 12% glycerin and be done. The starter just ensures they were viable to begin with. 2) It's totally possible to grow a 10million/ml inoculation rate in a 1L starter into 200B cells. Every strain is different, and has...

Yep, I put it into a cup of slightly warm water, and try to thaw it as fast as possible without a lot of heat. Just swirl it, etc, and try to get it to roughly the same temp as the starter as not to shock it. Yeah, the extra 5 ml is for expansion, or if I didn't eyeball the volume of the decant...

I think the problem is making the 2L starter from 27B cells only gets 2x more. Let me give an example of what I typically find for cell counts in my starters on a stir plate. Start with 100-150B cells in a smackpack or vial, add to 1L starter, and you might get 1.5x, maybe less. I find that an...

Negative or positive pressure is definitely not needed for doing this work, though access to that would significantly decrease the chance of any contamination. Positive pressure would be pretty easy, just put a high efficiency filter in front of a box fan blowing down into a plexiglass box with...

Yep, thanks. Typo there, 25% would be 25ml glycerin and 75ml water. Truth be told, anywhere from 10%-20% final glycerin concentration would give pretty similar results.

Glad I could help anyone out there, and thanks for the compliment!

You don't have to use a microscope for any of it, I guess that's why I left it out. Microscopes are fun, and to a point are useful in qualitatively measuring bacterial contamination in the brew (with good optics), but not necessary. For slants and plates, the colony from a single cell will be...

Any kind of alcohol would be fine, you could use some cheap vodka in a pinch. I've done the tubes straight into the freezer and the cells noticeably took a hit, the starters had a couple day lag time. You can also put the tubes in a styrofoam box in the freezer, and it has the same effect...

Thanks! All this information is out in the HBT space, but hopefully this condenses it a bit to make it easier for folks to start ranching.