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InvisibleInfiniteDreams
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Registered: 10/25/19
Posts: 1,224
There was an attempt (agar temp?)
    #26464837 - 02/02/20 08:33 AM (3 years, 11 months ago)

Armed with a PC and SAB I ventured into the world of agar for the first time.  Many months of reading and rereading the fantastic teks from our TCs and following threads on this site.  And it went quite well I thought, until the end.

Prepped the MEA recipe, dissolved in a boil and funneled into the media jars.  Then into the PC for a 20min sterilization session at 15PSI.  Followed bod's tek and I RTFM and that went well, although very time consuming.

But when I got to the pours it took a left turn, or two.

I got a fancy infrared thermometer I was hoping to take the guess work out.  When it read 117F I did the pours.  Then I let them sit for 5 minutes (planning to start a few germ plates same session).  Actually probably let them sit for 15 minutes.

And they weren't set yet!  I accidently spilled one checking it.  So either my thermo is way off calibration, or reading through the glass of the media bottle gives a lower temp value than the actual agar.  Frustrating as it was taking forever to cool.  I'm sure this gets easier with experience, but any ideas on the thermometer?  I thought I was making life easier by picking that up.

Also I guess due to the too high temp, I have insane amounts of condensation. When they really had set I went ahead and streaked 4 plates from print, and left 1 as a control plate.  The dark print seemed almost adhered to the foil, but I'm hoping since the spores are microscopic that I really did a good application even without the visibility.  I used an inoculation loop made from wire.

I tried the cup of warm water trick and it did absolutely nothing to clear up the condensation.

In the future I think first I"m going to have make pouring plates its own event so I'm not trying to do too much all at once while I learn.


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InvisibleDiego


Registered: 12/30/19
Posts: 346
Re: There was an attempt (agar temp?) [Re: InfiniteDreams]
    #26464853 - 02/02/20 08:47 AM (3 years, 11 months ago)

Try inverting your plates to help with condensation


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Some do it for the income
But we do it for the outcome
Some of us are active
While others just let their mouths run - Chali 2na & Jurassic 5


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OfflineA.k.aM
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Registered: 10/27/19
Posts: 16,782
Loc: Gaming the system
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Re: There was an attempt (agar temp?) [Re: Diego]
    #26465118 - 02/02/20 11:19 AM (3 years, 11 months ago)

They take a while to set, I’d give it a couple hours.

Especially if you stack them, the heat travels and keeps them warm longer.

For condensation usually leaving them unwrapped in the SAB for a few days helps.

Also with the thermo gun make sure to hold it like half an inch from the bottle. I was pouring at ā€œ120ā€ and having problems then I moved the gun closer and it went up to 140s


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LAGM2020


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InvisibleInfiniteDreams
Male

Registered: 10/25/19
Posts: 1,224
Re: There was an attempt (agar temp?) [Re: A.k.a]
    #26465162 - 02/02/20 12:01 PM (3 years, 11 months ago)

Thanks for the tips!  I'm planning to do another pour next week to get more practice and I'll try these out.

For the 12-hour update, here are the pics of my germ plates.  Because of the condensation overhead pics won't show anything, so these are from underneath with light above. 

I'm hoping that is sediment in the plates, although the agar was wasn't cloudy at all when I poured.  No pic of the control plate, but it has the same sediment.  I thought I was overzealous of sterile technique, but if it is contams already, then I guess I am losing my mind.

I used parafilm to wrap the plates.  In reality, all of these plates are the same color, seems my camera is playing with the colors.

The 2 B+ germ plates:



The 2 Ecuador germ plates:



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Invisibletramalot
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Registered: 12/13/19
Posts: 199
Loc: USA Flag
Re: There was an attempt (agar temp?) [Re: A.k.a]
    #26465167 - 02/02/20 12:05 PM (3 years, 11 months ago)

Infared gun's are somewhat subjective so try knocking it down 10f...


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It's the Journey I enjoy!


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InvisibleInfiniteDreams
Male

Registered: 10/25/19
Posts: 1,224
Re: There was an attempt (agar temp?) [Re: tramalot]
    #26465220 - 02/02/20 12:45 PM (3 years, 11 months ago)

So, I had about 15 more plates poured for extras and threw them in the fridge.  Just checked them and they are clear.  Same agar, so that is not sediment from the agar.

I will wait and see.  Would they contam so quickly?  I may need to do a lot better with inoculation...

I'd hope they are just spores, but I streaked the Z pattern and you can see whatever it is all over the place.


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