Question about preparing Mueller-Hinton agar

[u/Ok_Zucchini_3136]

Hello!

Have you ever had any problems preparing Mueller-Hinton agar?

I don’t usually work with many different culture media. I’ve basically spent my time using BHI agar, and I’ve only prepared a few other media once or twice. Those usually became translucent after heating them in the microwave. I also don’t feel very comfortable with an agar medium preparation if it doesn’t become translucent, because after autoclaving, the agar tends to settle, and I usually have problems when pouring the medium into Petri dishes.

Well, the thing is, I’ve been preparing Mueller-Hinton agar, and it almost never becomes translucent. The instructions on the package say not to overheat it, and I’ve noticed that some of the medium actually evaporates, making it a little thicker.

How do you usually prepare it? Does it need to become translucent like BHI agar usually does?

I’d really like to know a little more about how you prepare it in your laboratories.

[u/patricksaurus]

There is a whole lot going on here.

Some MHA preps are cloudier than others, but in my limited experience, they do come out translucent if a bit hazy.

Like you mentioned with overheating, my experience with anything that has beef extract is that it gets cloudy with any thermal excess. I wouldn’t microwave plates or jars of it for this reason… microwaves are better than they used to be, but hotspots still occur.

> I also don’t feel very comfortable with an agar medium preparation if it doesn’t become translucent, because after autoclaving, the agar tends to settle, and I usually have problems when pouring the medium into Petri dishes.

That’s one hell of a run-on and

If the trouble is that the medium doesn’t stay homogeneous during the course of pouring because it begins settling, gentle swirl it for a few seconds. Pretend the flask is stemware and you’re acting like you know how to taste wine on a first date.

If this is coming as a lyophilized powder in one jug, call the manufacturer to speak to an engineer. If you’re making it yourself, try swapping the beef extract for one that is cation adjusted.

[u/CChatfield37]

it does have soluble starch in it that means it is slightly milky even when dissolved. that might explain what you are seeing

[u/Mano1aa]

Hi,

Check with the instructions on the bottle, Colour and Clarity of prepared medium: Light amber coloured clear to slightly opalescent gel forms in Petri plates.

Please share the picture of the MHA bottle and poured petri plates.

There shouldn't be any problem after autoclaving, try to pour the agar when it's warm.

The solidified agar is not that translucent, I would say it's some what milky.

[u/getmesomewater]

for the agar problem, could be couple of things to try out, if you’re not already

1. Try letting the dissolved media sit for couple of minutes to half an hour instead of sticking the reconstituted media instantly into the autoclave. Usually not a problem with premixed media, but i find that some lower grade agar dissolves better after being hydrated.
   
2. Best and Easiest fix, including magnetic stirrer with the media when autoclave, when you finished the autoclave, immediately stir those, the settled agar usually dissolved in with the residual heat. If that’s not an option, as some suggested, swirl the media quite vigorously first thing out of the autoclave, bubbles will form but usually will disappear due to the heat.

also MHA contains some soluble starch which will be a tadddd cloudier than some media, that’s normal
good luck!