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Offlinecatfeesh
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Failure = Glove box idea
    #9627366 - 01/17/09 04:59 PM (15 years, 2 months ago)

Hello everyone, awesome website, the avatars here are the best.

My Question:
I was wondering if anyone had devised a glove box where one could poke the syringe out through the glove box into an alcohol flame sitting outside the glove box?

Poking the syringe through a patch of tyvek, or medical tape into a burning flame for sterilization would allow total containment of the glovebox...the only air transfer into the glovebox would then be through the aforementioned tyvek window.

The needle could be pulled back through the window while red hot, or even wiped down with alcohol afterwards from within.

The reason I ask is my first inoculation failed totally...contamination in all 10 jars.
The Trichoderma? contamination looked as if it had sprouted from the very points of my inoculations.  Id love to blame the spore provider, but I'll chalk it up to being a noob.

I watched RRs videos a few times and followed the PF Tek to the word.
The weak link in the PFTek process seems to be flaming the syringe outside of the glove box, allowing contaminated air into the sterile work area.

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Offlinedead
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627375 - 01/17/09 05:00 PM (15 years, 2 months ago)

That's an interesting idea, but instead of tyvek I would make a small injection port in the side of the glovebox.


--------------------
"The third eye. You spend years doing everything you can to open it and then the damn thing opens and your friends laugh at you when you tell them you can see their souls behind their eyes burning like rainbows."

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InvisibleShroominit
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627377 - 01/17/09 05:00 PM (15 years, 2 months ago)

My guess is that you used a contam'd LC.

I did that before.

I think the alc flame would have to be too close to the GB to be effective.

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Offlinedead
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Re: Failure = Glove box idea [Re: Shroominit]
    #9627394 - 01/17/09 05:03 PM (15 years, 2 months ago)

Just refrain from using any flammable desinfectant inside the glovebox and you can use a lighter inside it.


--------------------
"The third eye. You spend years doing everything you can to open it and then the damn thing opens and your friends laugh at you when you tell them you can see their souls behind their eyes burning like rainbows."

Links:
:regularshroom:Nibin's Guide for Noobs
:regularshroom:some easy teks on bulk & grain prep. (my journal)

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Offlineexzile
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Re: Failure = Glove box idea [Re: dead]
    #9627414 - 01/17/09 05:06 PM (15 years, 2 months ago)

I JUST use the kitchen stove........
and inoculate my jars right there on the stove...
no failure yet.


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InvisibleShroominit
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Re: Failure = Glove box idea [Re: exzile]
    #9627426 - 01/17/09 05:07 PM (15 years, 2 months ago)

Quote:

exzile said:
I JUST use the kitchen stove........
and inoculate my jars right there on the stove...
no failure yet.




If you have a self-sealing injection port and flame the needle/alcohol wipe the port, you're pretty much good no matter where you do it.

Also, I'd recommend against using flames in any enclosed area. Especially a plastic one. Also I use alcohol for mine, so it wouldn't go to well.

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Invisiblearchivist
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627444 - 01/17/09 05:10 PM (15 years, 2 months ago)

I do something similar to this with my glovebox, except I don't put any sort of filter material in the hole.  I just make the hole a little bit larger than the diameter of the syringe.

I don't think the Tyvek idea would work because the red hot syringe would surely burn the Tyvek when you're withdrawing it.  It hasn't been necessary in my albeit limited experience as I've yet to experience a contamination in a jar so far ::knock on wood::.

If you had a 100% contam rate, odds are more than likely your spore/LC syringe was already funky.  Did you order from a sponsor?


--------------------


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Offlinecatfeesh
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Re: Failure = Glove box idea [Re: archivist]
    #9627633 - 01/17/09 05:44 PM (15 years, 2 months ago)

Archivist: The port idea is cool...I'm going to try it.

I probably should have alcohol wiped my medical taped injection ports, prior to inoculation but it's hard for me to believe they were contaminated as they were protected by the tin foil...totally possible though.

Yeah, I did order from a reputable sponsor.  I don't know if I should bug 'em about it.

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InvisibleShroominit
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627690 - 01/17/09 06:00 PM (15 years, 2 months ago)

Quote:

catfeesh said:
Archivist: The port idea is cool...I'm going to try it.

I probably should have alcohol wiped my medical taped injection ports, prior to inoculation but it's hard for me to believe they were contaminated as they were protected by the tin foil...totally possible though.

Yeah, I did order from a reputable sponsor.  I don't know if I should bug 'em about it.




Did the micropore tape get wet when PCing? Wet tape pretty much = contams because it holds moisture so well. Regardless, if it wasn't an LC syringe (which if it's active, it would not be), then its probably not contam'd.

Try putting the jars in a GB to cool (after PC) and then innoc 12-24 hours later, still in the GB. Try not to use too much spore solution and definitely flame the needle, wipe the tape with alcohol and innoc quickly. Then replace tape.  If this procedure fails (with trich again) email the sponsor, they may rectify the situation.

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Offlinecatfeesh
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627801 - 01/17/09 06:24 PM (15 years, 2 months ago)

Shroominit, are you recommending 2 layers of tape, or just replacing the punctured tape with a new piece?

Yep, the tape was pretty wet, and I was using spore syringes (Brazilian strain).
At the time of inoculation I was thinking about lifting the tape off to inoculate, but figured it wouldn't stick down again due to the wetness...I figured why use tape if it has a hole in it?

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OfflineABC
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627811 - 01/17/09 06:27 PM (15 years, 2 months ago)

Personally, I use a still air box because I can take my hands out and flame the needle that way.

Needles are relatively short, I might be worried about having the plastic tote melt or catch fire if you had to flame the needle through the side :shrug:

edit: welcome to the shroomery!

Edited by ABC (01/17/09 06:29 PM)

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InvisibleShroominit
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627829 - 01/17/09 06:30 PM (15 years, 2 months ago)

Depends. If you have small holes like me, needle sized holes, then you just tape over it. It's fast and it works.

If you have 1/4" holes or something then you probably don't wanna double up on tape as it will greatly restrict GE.

Wetness is EVIL when talking about tape. It will allow contams to permeate through the barrier that it creates.. and then contam your jars.

ABC, that's exactly what I was saying. Even with 2.5" or 3" needles, you can't flame the whole thing through the side of the GB, just take it out and bring it back in. If you're not doing G2G transfers at the same time, you'll be Ok.

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Offlinemudkip
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Re: Failure = Glove box idea [Re: Shroominit]
    #9627879 - 01/17/09 06:41 PM (15 years, 2 months ago)

I would think if your using PF jars you should have a verm layer, and tape it after inoculation.  And I doubt all his jars contamed from wet tape.  I'd say its the spores based on information given about the contaminations starting at inoculation point. 

Also, Your glovebox does not need to be airtight, I know people who use a fish tank upside down, moved off the table enough to put hands in.  It's not so much about keeping contaminated air out, its about the air being still so there is not bacteria blowing around.  Also keep in mind the needle should not touch anything after its been flamed.  Did you flame in between jars?

I know its not recommended but I put an alcohol soaked cotton ball on around the needle after flaming, this helps me because i can hold the needle in place better (I have shakey hands somtimes)  and contams cannot enter (in theory anyways).  Seems to work for me.


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InvisibleShroominit
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Re: Failure = Glove box idea [Re: mudkip]
    #9627883 - 01/17/09 06:43 PM (15 years, 2 months ago)

Quote:

mudkip said:
I would think if your using PF jars you should have a verm layer, and tape it after inoculation.  And I doubt all his jars contamed from wet tape.  I'd say its the spores based on information given about the contaminations starting at inoculation point. 

Also, Your glovebox does not need to be airtight, I know people who use a fish tank upside down, moved off the table enough to put hands in.  It's not so much about keeping contaminated air out, its about the air being still so there is not bacteria blowing around.  Also keep in mind the needle should not touch anything after its been flamed.  Did you flame in between jars?

I know its not recommended but I put an alcohol soaked cotton ball on around the needle after flaming, this helps me because i can hold the needle in place better (I have shakey hands somtimes)  and contams cannot enter (in theory anyways).  Seems to work for me.




I didn't say they all contaminated due to wet tape, I was just saying that wet tape is definitely an entry point for contams. I mentioned that the innoculant (be it LC or spores) could be contaminated, more likely with Lc, of course.

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Offlinecatfeesh
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627895 - 01/17/09 06:46 PM (15 years, 2 months ago)

Alright, I'll be extra vigilant next time - thanks for the good advice, I'll get these beasts sprouting eventually :thumbup:

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Offlinemudkip
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Re: Failure = Glove box idea [Re: Shroominit]
    #9627936 - 01/17/09 06:56 PM (15 years, 2 months ago)

Quote:

Shroominit said:

I didn't say they all contaminated due to wet tape, I was just saying that wet tape is definitely an entry point for contams. I mentioned that the innoculant (be it LC or spores) could be contaminated, more likely with Lc, of course.




Oh no man I didnt mean it like that, I was just basically trying to say he should probably leave the holes open with the tin foil on during sterilization. 

I personally kick it with glass drinking cups, tin foil as a lid haha.


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Invisiblenw_shroomy
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Re: Failure = Glove box idea [Re: Shroominit]
    #9627950 - 01/17/09 06:59 PM (15 years, 2 months ago)

we are talking PF jars here.There is a dry verm layer so IME it dont matter if his tape got wet if the dry verm layer is still DRY.My tape always gets wet right before I inoculate as a couple of drops always comes out of the needle anyway.Also if he followed RR's video to a T like he says then he kept the tinfoil on until he was ready to inoculate. I remove the tinfoil in the glove box.
I would say that it was a bad syringe


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Offlinecatfeesh
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Re: Failure = Glove box idea [Re: catfeesh]
    #9627993 - 01/17/09 07:08 PM (15 years, 2 months ago)

I did flame between jars, but maybe I should have been more aggresive - I was a little wary of melting the syringe.  I tried to follow the PF tek, and RR's vids as closely as possible.
I had a verm layer, but I'll try taping AFTER inoculation next time, so the tape will be dry.

Is spore syringe contamination common? 
I should have taken photos of my jars, because the contamination matched exactly where the inoculant flowed into the substrate.  Bizarre.  I'm going to try PC the metal needles next time.

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OfflinePsychOfMSE
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Re: Failure = Glove box idea [Re: catfeesh]
    #9628013 - 01/17/09 07:11 PM (15 years, 2 months ago)

Quote:

catfeesh said:
I should have taken photos of my jars, because the contamination matched exactly where the inoculant flowed into the substrate.  Bizarre.  I'm going to try PC the metal needles next time.




Not bizarre.  Just means you have a contaminated syringe.

Good luck!


--------------------
"I think it's impossible to really understand somebody, what they want, what they believe, and not love them the way they love themselves." - Orson Scott Card

Agar's Grain LC Tek <--- Never fool with honey water and contaminated LCs again!
Large_Dose's Ohmatic Monotub Tek
Monstermitch's LC/WBS/Monotub/Casing Tek

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InvisibleShroominit
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Re: Failure = Glove box idea [Re: PsychOfMSE]
    #9628034 - 01/17/09 07:14 PM (15 years, 2 months ago)

You could take pics and contact vendor.  They'll probably ship you another.
The only reason I encourage testing is because most people who have contamination problems are just starting out and don't get sterile procedure down well enough. Plus its too easy to just blame the vendor all the time.

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