My frozen yeast died

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I froze some yeast (blend of US-05 and whatever Odell uses) a few months ago, and followed the instructions in the article on HBT exactly. The day before brewing, I made a starter with the yeast, and it didn't seem very active. Nonetheless, there was a layer of yeast at the bottom of the jug the next day, so I assumed it was okay. I pitched the yeast as usual, at 65F. 24 hours later, and there is zero activity. No krausen, no airlock movement. No bubbles or things moving around in the wort. For this yeast, this isn't normal. When I used it the first time, it took off after a few hours. I went ahead and pitched some wyeast 1007 that I had in a mason jar in the fridge. I figure that more yeast is better than sluggish and/or dead yeast and risk of infection.

Has anyone else had bad luck freezing yeast? One thing I noticed, the yeast in the freezer turned somewhat darker and didn't look as "creamy" as it once did. Was this a bad sign?

 

[kh54s10]

How much yeast was in your frozen sample? I make 20ml vials with 5 ml yeast, 5 ml glycerin and 10 ml water, chill overnight in the fridge then freeze. The vials are in a foam box with some ice packs. Other ice packs are on the outside to counteract the defrost cycle of the freezer.

Since I am starting with only 5ml yeast I make a weak 250 ml starter wort and let that go for 24 hours then I make a .5 liter addition for another 24 hours then I figure I have about the same amount of yeast as a White Labs vial or Wyeast Smackpack. I then make a proper sized starter. I run it for 24 hours then chill to decant.

So it takes almost a week to make a proper starter from my frozen yeast.

I suspect that either you pitched very little yeast, or it had been thawed then refrozen, harming or killing most of the cells.

The darker color doesn't sound right to me but I don't really know. Mine always look the same. I have successfully revived yeast stored for more than 2 years.

 

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A week? Maybe I needed more time. The article recommended using one or two vials for an overnight starter. I used 25 mL yeast slurry and 25 mL 20% glycerol solution in a 50 mL vial. I stored the vials in super saturated NaCl solution since I have a defrosting freezer. The article recommended using isopropanol, but I didn't like the idea of open isopropanol hanging out with my frozen foods. Someone in the comments recommended using super saturated brine as an alternative.

 

[kh54s10]

A fresh White Labs vial of yeast is said to contain 35 ml yeast. I would suspect that your 25 ml frozen sample was probably a bit less due to less than laboratory conditions.

Mrmalty, for a 1.055 ale suggests a 1 liter starter with one vial. Since the frozen yeast have been dormant for quite a while I would expect a larger starter and give it more time on the stirplate.

As I said I am starting from only 5 ml yeast so I have to really work at building cell counts.
I have always gotten action by the next morning after pitching in the early evening.