Background
I’ve been doing tissue culture work on different plants and had constructed a sterile glove box in which I’d be able to work in an environment free of organisms that could contaminate the cultures.
The concept is that the tissue/explants are placed inside the box and the UVC germicidal light turned on once the lid is locked shut. This will eliminate the contaminants (bacteria, viruses, mold) in the inside air and create a seal that won’t allow the life-filled outside air in. I shut the UVC light off after 3-4 minutes of being on before working, as the light will damage my eyes. I then sterilize the surface of the explants for clean plating, allowing for a successful callus induction.
Here’s what I’ve come up with…
The Problem
Some of the cultures have been becoming contaminated by mold spores. Just for fun I let them run wild for over a month…
So what’s happening here?
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The seal created by the gasket in the lid is leaking. This is caused by working with the gloves creating both negative and positive pressure leading to outside air entering the box.
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The germicidal UVC light isn’t killing everything.
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My sterilization technique for the explants needs to be improved.
My guess is that it’s either or both of the first two. I know for sure that the light works to some extent cause I can smell ozone, and I know of the leak cause I can smell it once I begin working but not before I start. I can determine if it’s also the third by eliminating the first two.
The Solution
Convert it into a filtered positive pressure system that will constantly blow sterile air into the box and through an exhaust port on the bottom. Even if there’s a leak, nothing can get in cause filtered air is always being pushed out.
I will be keeping this thread updated as the project progresses… Feel free to coment and ask questions! Cheers! 
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Have you thought about trying to make a simple laminar flow hood to work in front of?
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Yes, I’ve looked into it. To build a more professional type of laminar flow hood would be a lot more costly and take up a bit of space. Not to mention is not portable like this one is. I eventually want to build a nice one but I plan on moving in the future and want to be able to take it with me.
I also want to show that one can be constructed with fairly common supplies for aound $100.
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If your going to try for that. Maybe set up an exhaust fan for the chamber nd pull through like a merv 12 or 13 filter. That should catch basically all contaminates.
Where did you get your fittings for the gloves? I like those.
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Sorry to hear that you’re having issues but I really like the build - it looks really clean.
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Looks similar to something I used for inoculating jars for mushroom spores.
If you are stretching the gloves and create air movement you could try extending them with flexible ducting tot try to lessen the air seeping through the lid seal. Just a thought and not sure if it would solve the issue.
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The idea I’m working with is to pull air through a particulate filter rated for biological contaminants before sending it through a fan.
These are 3 types of filters designed for military gas masks or respirators…
The one on the left is a 1982 filter for a Russian GP-5 mask. I’ll explain later what this will be used for. The middle is an Israeli Supergum gask mask filter manufactured last year, rated for NBC (Nuclear, Biological, and Chemical protection.) I won’t be using this one but thought I’d show it anyway. The last is a particulate filter approved for bio made in the same factory. If it’s good enough for the Israeli military and civilian medical use, it’s good enough for me. Costed $20.
This is the Russian millitary surplus GP-5 mask kit. The filter likely contains asbestos so don’t breath through it! They claimed to stop using it after '79 but that’s coming from the Soviets so take it with a grain of salt…
I’m going to use an air mattress pump as the fan system. Attached a rheostat/fan speed controller to slow it down. I want a steady consistent flow of air, not a hurricane…
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4-Inch duct flanges from Woodcraft…WoodRiver Dust Connection 4" Flanged Inlet Fitting for Dust Collection System | Woodcraft
Attached the gloves to them with plumbing straps from the local hardware store.
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@Foreigner Thanks for the kind words!
@MBVapester This can be applied to mycology, or any situation where you need a clean working environment to do cultures in.
I thought about just trying to stop the leak altogether but doing this may even allow me to just remove the gloves making it even easier. My main concern is that the air won’t flow evenly and create vortexes where spores will get caught and just spin around in.
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Are you going to cut a hole in the side and draw through the gas mask filter, blowing into the tub?
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I’ll be blowing from the top, you’ll see what I have planned. I’m waiting for my 3M respirator filters to arrive so I can safely cut the Russian filter without inhaling anything dangerous. 2020 taught me all about filters and respirators/masks.
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I think your causing more issues having a fan. I use a SAB with no gloves attached. Disinfect the room with iso before you get to work. Spray inside of box also and clean each time. The physics of how the air will drop once it reaches inside the holes is why it works.
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Thanks for responding. I’ve tried spraying the inside of the SAB with a diluted beach solution and still, there are issues. I’ve heard of people having success with an aquarium flipped on its side. This would make sense with what you’re saying as the air should fall downwards. So, maybe removing the gloves altogether will solve it?
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And people say smoking pot fries your brain and makes you lazy don’t know where they get that info from just reading posts here shows some dedicated work and thought put into some of these projects
Peace and stay safe
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This is where I got my info.
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hey in orchid propagation they use hydrogen peroxide solution to make it sterile. Ever tried that? I would read up on orchid flasking, it contains some useful info most likely applicable.
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I lysol all around my room, then paper towels dampened with h202 on the bottom of the SAB to get any falling spores to stick and not float back up. And run a fan with a filter on its back to catch spores for an hour before. Turn off when you lysol.
I finally picked up some filters and made 2 again.
I use one in the house for dust too.
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I use a fish tank on its side for flasking.
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Hi. Did you pressure cook all your instruments or heat sterilise them?
You should always clean all surfaces with 75 or higher proof alcohol. This includes gloves the box etc. Always work in small area with little air circulation to avoid airborne particles, mold spores like the jar on the left (Trichoderma or penicillium molds)
You can always use HEPA filters for vacuum cleaners… but you can work in asepsis by using a Bunsen burner or an alcohol lamp.
Nice work. I like the box 
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