Preparing sterile petri dishes?

[Paul Anderson]

I've finally picked up some nutrient agar mix and petri dishes. How do I prepare it, pour it and keep things sterile at the same time? After preparing the agar, can it be remelted? Some sites imply this but don't explicitly indicate either way.

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[Eric Ma]

Hi Paul,

Yes, I have made LB-Agar, autoclaved it and have it solidified, only to find that I need to pour a sleeve of plates later, so at that point I would melt the agar in the microwave and pour them. The only thing that happens is that the agar might change color (not sure why, my guess is that some chemical rxn not unlike what happens when you fry noodles on a pan), but that doesn't seem to affect the growth of my bacteria.

Hence, my answer is: go ahead and re-melt your agar!

Cheers,
Eric
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On Fri, Nov 5, 2010 at 7:43 PM, Paul Anderson <wacky...@gmail.com> wrote:

I've finally picked up some nutrient agar mix and petri dishes. How do I prepare it, pour it and keep things sterile at the same time? After preparing the agar, can it be remelted? Some sites imply this but don't explicitly indicate either way.

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[Eric Ma]

On another note, to pour it in a sterile fashion, make sure you do it in a biosafety cabinet or near to a flame. I prefer the BSC because it's easier to guarantee sterility, but then again it also is quite a bit of a hassle to ethanol everything before putting things in the BSC.

Cheers,
Eric
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On Fri, Nov 5, 2010 at 7:43 PM, Paul Anderson <wacky...@gmail.com> wrote:

I've finally picked up some nutrient agar mix and petri dishes. How do I prepare it, pour it and keep things sterile at the same time? After preparing the agar, can it be remelted? Some sites imply this but don't explicitly indicate either way.

Sent from my iPhone

[Cathal Garvey]

As long as the plates are already sterile you can pour near a bunsen flame when the temperature cools to 'uncomfortable but not burning'. If you want to add antibiotics, wait a bit longer til you can hold it with your hand: what won't hurt you, won't hurt antibiotics.

If you've added antibiotics, pour it all immediately or you'll be wasting some, because remelting agar requires near-boiling temperature which will degrade antibiotics.

Autoclaving media causes browning reactions, same as frying or grilling makes things go brown. A little bit is fine, but repeated Autoclaving and you might affect your results. Does burned LB lead to faster or slower growth? Poor buffering? I don't know. General advice is just to pour it all or keep it at 50C til you need it (at this temp it won't gel).

Remember, you only need enough to cover the plate surface, thick plates are usually unnecessary. You can get way more plates through frugal pouring!

On 6 Nov 2010 02:42, "Paul Anderson" <wacky...@gmail.com> wrote:

I've finally picked up some nutrient agar mix and petri dishes. How do I prepare it, pour it and keep things sterile at the same time? After preparing the agar, can it be remelted? Some sites imply this but don't explicitly indicate either way.

Sent from my iPhone

[Paul Anderson]

On Sat, Nov 6, 2010 at 4:01 AM, Cathal Garvey <cathal...@gmail.com> wrote:
>
>
> Remember, you only need enough to cover the plate surface, thick plates are
> usually unnecessary. You can get way more plates through frugal pouring!
>
>

I have some small disposable plastic petri dishes. These should
withstand a pseudo-autoclaving in a pressure cooker, right? The media
I've bought is LB nutrient agar. Seems like it makes about a liter.
I've got a liter glass bottle I'll be storing it in. So I will mix up
the media(they say to add one liter of water to the packet contents,
then autoclave), pour into the bottle. Does it need to be boiled to
make it set? Or is cooling sufficient?

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[Nathan McCorkle]

Its basically the same as making jello... add the stuff that can be
cooked to water in right amounts, put in glass jar, and autoclave...
maybe slightly warm and mix everything prior to autoclaving, just to
be sure it all dissolves properly (stuff that can't be cooked are some
amino acids, when cooked the polymerize so they're not free AAs
anymore... and a lot of drugs, these break down with heat, usually you
can look up this temp and figure out if they're autoclave safe)

after autoclaving then can you pour into petri dishes... the media
will spill if you try to autoclave petris, and plastic petris should
be one-use only.

if it solidifies in the bottle, use a microwave in 30 second
increments to re-liquify... you want to avoid superheating and
boil-over... as well you want to slowly raise the temperature so you
don't over-cook the media or change the concentration by evaporating
water too much. then let the media cool a bit while still in the
bottle, it makes handling easier and avoids breaking down any
chemicals you wish to add.

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[Paul Anderson]

Can the plastic petri dishes be autoclaved? If not, how should I sterilize them?

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[Jacob Shiach]

If the plastics not autoclaveable you may be able to get away with using a dilution of bleach.

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[Nathan McCorkle]

You can try it, but they might deform and be more susceptible to
evaporation of cell media or air contamination. For most stuff, other
than genetic selection, you can usually do without
antibiotics/antifungals, as long as you keep everything sterile.
Aren't the plastic plates sterile, or even in a sealed plastic tube?

For Convenience:
http://www.onlinesciencemall.com/Shop/Control/Product/fp/SFV/30852/vpid/7531985/vpcsid/0/rid/126318

For those that want to autoclave:
http://www.onlinesciencemall.com/Shop/Control/Product/fp/SFV/30852/vpid/9129502/vpcsid/0/rid/126318

wide-mouth half-pint mason jars also work great for petris... a little
trickier to see your cultures though... free shipping to an ace store:
http://www.acehardware.com/product/index.jsp?productId=3376864&kw=wide+mason&origkw=wide+mason&searchId=49816940464

if someone wanted to get something in-between, a case of petris and
two cases of half-pints would get you top viewability. I just measured
a canning jar to be 86mm across, so the 100mm petris should fit
alright.

If you want to re-sterilize plastic, I've previously soaked them with
70% ethanol spray and let dry in a HEPA flow-hood. Should work
similarly with isopropanol.

[Aaron Hicks]

Disposable petri plates will deform and melt under autoclave conditions (which, as the autoclaves I've used run at 15 psig, would be the same pressure- and, therefore, temperature- at which a pressure cooker used to sterilize media would run at). They're just water-clear styrene, and PS has a glass transition temperature of 95C.

Buy 'em sterile, pre-nuked at some hideous concentration of radiation.

-AJ

[Brian Degger]

Otherwise you can get a few glass petri's and they are fully reusable.

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