FlytrapCare Carnivorous Plant Forums

Hey all,

I recently built myself a glove box to do some tissue culture work in, and I have been debating on how I am going to make sure the inside is sterile once I close it up. I have been thinking of taking all of the hot sterile water and media jars, and putting them inside on the silicone mat, closing up the box with a spray bottle of a sterilizing agent inside, and then spraying it down. I'd then let the media and water cool down in the sterile glove box, and would come back to work when everything is room temperature, and possibly spray again if needed.

Which do you think would work best as a sterilizing agent for the whole inside of the box:
-Bleach 10-30%
-H2O2 ~3% (storebought)
-Isopropyl Alcohol 91%
-White Vinegar and then H2O2 (http://cleaningbusinesstoday.com/blog/h ... ecting-duo)
-Premixed PAA (using white vinegar and h2o2)
-Other

My glovebox is a plastic storage container that I modified, with some pvc pipe inserts, so the chemicals can't be able to damage any of that. If any of the above chemicals are known to damage plastics, please let me know.

What about baby disinfectant ?
We have a stuff in the UK called Miltons.

http://www.boots.com/en/Milton-Sterlisi ... 20-%20Baby

"Milton Sterilising Fluid allows you to sterilise in 15 minutes only, killing all bacteria, viruses, fungi and spores (tough dormant bacteria). It has been used in hospitals for many years as a simple and very reliable method."

-Isopropyl Alcohol 91%

The only thing that would be better than that would be ethanol alchohol. But that is more expensive and harder to get in high concentration (it's moonshine).

As for the concentration, I've heard 70% (either isapropanol or ethanol) actually works better as a disinfectant than 90%+ concentrations. It's easy to dilute a bit with water though.

The only thing you have to be careful about is your glove box. Some plastics don't react well to alcohol and will crack. You'll have to see if the type of plastic the shoe organizer is made out of handles the alcohol or not without degrading. If it's acrylic (plexiglass) you shouldn't use alcohol.

The box should have a little triangle of arrows with a number molded into it somewhere, it's a recycling code. If you look up the number in the triangle that will tell you what kind of plastic it's made out of.

Household antibacterial wipes are quite effective in the cleansing and sterilization of surfaces. I generally avoid any sprays or aerosols, anywhere near a hood . . .

@Jaws... actuually have some Milton tablets I am planning on using for some explant sterilization. I'm not sure if it can be used as a spray, or if you need to soak things to sterilize them though.

@entroypy81... Thanks, If it can handle the alcohol, I'd love that, because it dries without any residue, and is easy to obtain.

@BigBella... Why don't you use sprays near hoods? Is it because of the filters? I don't have any airflow with this one, or filters/fans, so I figured a spray would knock any airborne particles that are there once I close it up, onto the sides.

You can buy Miltons as a spray, but id say make the solution up from your tablets and put in a sprayer.

Are you a bugs nightmare?


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Doomsday wrote: @BigBella... Why don't you use sprays near hoods? Is it because of the filters? I don't have any airflow with this one, or filters/fans, so I figured a spray would knock any airborne particles that are there once I close it up, onto the sides.

Initially, I was referring to a hood and hepafilter arrangement; but also because most effective antibacterial solutions / sprays are very irritating to the eyes and mucous membranes -- especially in a closed, poorly ventilated setting where TC is most likely to be performed by hobbyists.

I have come to use the wipes in cleaning surfaces and also as a rest for lids while I am either establishing or re-plating cultures and don't wish to juggle lids, along with bottles, in the same hand. I also wipe bottles before applying Parafilm; and my contamination rates remain below 1% . . .

@Jaws... I haven't used any of my tablets yet, but I have enough to make a decent amount of solution to last a while.

@bijillon... Yup, that's me!

@BigBella... Yea, I definitely noticed that bleach and alcohol were irritating when I used to use them in my fishtank (laying on it's side), and hated the smell, but figured it might not be an issue this time, since the box is completely sealed up.
I like the idea and ease of the wipes, I just think it might be hard to get the entire surface wiped down once it's closed up. I could wipe it before closing it, but that might not end up being so sterile.
I also like the idea of using them to place lids on while working in the jars. Thanks.

Why not quaternary ammonia? It will affect bacterial spores where alcohol will not and does not have the same potential to cause breathing issues in small spaces as does bleach. QA does need more contact time than bleach (~15 minutes). CDC info says that 70-90% alcohol work equally as well against non-spore forming bacteria. 90% is more flammable and evaporates more quickly. Both are ineffective against bacterial spores.

TCGator wrote:Why not quaternary ammonia? It will affect bacterial spores where alcohol will not and does not have the same potential to cause breathing issues in small spaces as does bleach. QA does need more contact time than bleach (~15 minutes). CDC info says that 70-90% alcohol work equally as well against non-spore forming bacteria. 90% is more flammable and evaporates more quickly. Both are ineffective against bacterial spores.

Have you ever spent some quality time in closed quarters with either Simple Green or Microban? Damn! Clorox Broad Spectrum Quat is about the least offensive or yack-worthy; and we have used that for some general disinfection.

It should be mentioned that the cleaning is also a mechanical effort, as in the washing of one's hands -- and not necessarily ensuring the elimation of all potential spores. In our experience, a decent, thorough mechanical cleaning yields very little on streak plate cultures that we've used for periodic testing . . .

I'm thinking about giving everything going into the box, including tools, plates, jars, and the box itself, a wipe down with antibacterial Clorox wipes, and then about 15 minutes before and then again right before working, giving a light spray of 70% alcohol from the inside, to knock any airborne particles out and onto the walls of the box.

The iso shouldn't be a problem since I have no air filtration system or air current of any kind, and I also have the box closed up, so very little of the fumes would be able to get out, and I doubt it'll be very irritating. It also dries quickly and leaves no annoying residue to clean off.


As for my tweezers and scalpels... If I wipe them with Clorox wipes, and then sit them on something sterile, will the residue affect my explants/cultures once it's dry? If so, would you recommend a dip in sterile water, or h2o2, to remove the bleach residue?

I ended up using Lysol antibacterial wipes ("kills 99.9%... I") on everything going into the box. I finished with a spray of diluted iso alcohol once the box was closed up, to clear the air of any floating particles.

I flasked up some seeds and two dionaea explants... No visible contamination yet, (I flasked everything up on Monday).

So, the flasking process appears to have been pretty sterile. Springtails crawled into three of the jars, though. One each, but they still ruined them, obviously. I swapped the saran wrap for micropore tape and moved the other jars away from my plants. They are still sterile.