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OfflinePsilocyBen17
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Sterile Technique Discussion Thread * 1
    #23501482 - 08/02/16 11:14 AM (7 years, 5 months ago)

Sterile Technique Discussion Thread

Developing good sterile technique is one of the most challenging (and certainly the most frustrating) parts of mushroom cultivation. I've noticed lately that its easy to find 100's of threads stressing the importance of good sterile technique, but much harder to find threads on how to develop it. I find threads giving line by line instructions (such as The Tiger Drop) incredibly valuable, especially for newcomers.

After 2 some years in this hobby I have advanced from open air innoculations in a bathroom (:facepalm3:) to a still air box, to a flow hood. Currently I'm happy with my success rate of about 94%, but there is always room to improve our understanding and technique. There are still some things I don't understand. I'll pose some questions for discussion and encourage others to do the same

Although Still Air Box vs. Flow Hood discussions may be fun lets move forward with the understanding that a flow hood is not a magic wand....it does not cast a spell on the cultivator protecting him or her from future contaminates. A SAB and A FH offer the exact same chances of success to the cultivator. A FH  offers more comfort than a SAB, but they are interchangeable tools in terms of technique. I hate when newbies dream of flowhoods or blame their high contam rate on being confined to a SAB. A flowhood will not correct your mistakes or refine your technique. That part is up to you.

The questions I've been pondering about technique lately are as follows:

1. Preperation: what do you do?

Most days I will spend almost an hour prepping the work room before I begin work. I'll  put my hood on the lowest speed and wipe down the work space and bench with iso (or household cleaner and then iso if it really dirty). Then I will lay out all my jars or plates for inoculating and wipe them down too., especially the rim of jar lids. Next, I'll shower very thoroughly, shampooing my hair, scrubbing my arms and legs, pits, and all other parts. I'll towel off with a clean towel, brush my teeth, and gargle with mouthwash. I don't change into clean clothes because I do all my work in the nude.  I know my skin is cleaner than anything I can put over top of it. :shrug: Plus i think its important to feel vulnerable and in a precautious state of mind while in front of the hood. Next I'll put on my mask and gloves if I decide to wear them, wiping down my hands and forearms with iso,  and take a comfortable seat infront of the hood. Meanwhile the hood has been running and the door has been shit so I know the air in the room is clean....If I'm doing any sort of tranfering from agar I'll wipe down my scalpel again and do a wipe and flame inbetween transfers using a microjet blowtorch. All my transfers/innoculations happen as close to the hood as possible and of course always happen upstream.

Although the prep sounds like a pain its really not....i think its the part of cultivating that I enjoy most. I like being meticulous and paying attention to detail. What frustrates me though is that its often hard to see the payoffs of good preparation....somedays, if I'm in a rush, I'll skip having a shower completely and instead will just give my hands a good wipe with iso.... but I often won't see much a change in my success rate.....what do others have to say about this?

2. Rubber Gloves: Are they worth it?

Because I spend a lot of time outdoors my hands are constantly covered with little nicks and scrapes. I'm also a nail biter. Sometimes I look at my hands and to me they seem like a bacterial haven to me. When I started FH work I used to wear rubber gloves, but I find them incredibly hot and cumbersome. I found with gloves I would drop my scalpel alot and sometimes struggle to open lids. Nowadays I've stopped wearing gloves but will give my nails a good scrub in the shower beforehand and wash my hands and forearms very well.

I'm of the opinion that one of the cornerstones of good technique is a dexterity and a quick pace. Gloves prevent this.

3. isopropyl alcohol

I use 70% iso to wipe down my work area, tools, jars or plates before beginning sterile work. I will also use it to wipe down my scalpel blade prior to flaming it in between transfers. I can't help thinking that this may be overkill, especially since I learned Cron doesn't use iso that much because of the belief that it simply spreads contams around....what do others think? If I can cutdown on waste I generate during sterile work(iso, paper towel) I'd really like to. I find I'll use fresh papertowels to wipe down my jars after i've cleaned off my bench and workspace....is this necessary?

If my jars are hot out of the PC and have cooled infront of the air stream I won't bother to wipe them, but this usually never happens.  Seeing as I'm able to do 10 jars at once in my PC it weighs a ton when its full. Its too cumbersome to bring upstairs to my work room that easily so I usually just bring the jars up in a flat.

4. Cleaning the work room:

I always wipe and clean down my bench but I almost never vacuum or mop the floors because that room also serves as a spare bedroom and is very cramped with not much floor space.

Reading GGMM really scared me because Stamets cleans his lab daily and implies that working near a bed or any sort of fabric (drapes, laundry, bedding) will result in certain failure. I seem to be doing fine though. Whats the deal with that?

I know this post is long, but this is whats been on my mind :shrug: looking forward to some good discussion.

Pb


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OfflinePsykronik
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Re: Sterile Technique Discussion Thread [Re: PsilocyBen17]
    #23501526 - 08/02/16 11:25 AM (7 years, 5 months ago)

Nice write up  :asianofapproval:


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Invisibleweetsie
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Re: Sterile Technique Discussion Thread [Re: PsilocyBen17]
    #23501810 - 08/02/16 12:56 PM (7 years, 5 months ago)

Quote:

PsilocyBen17 said:

I do all my work in the nude.  I know my skin is cleaner than anything I can put over top of it. :shrug: Plus i think its important to feel vulnerable and in a precautious state of mind while in front of the hood.




:whoak:


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OfflinePsykronik
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Re: Sterile Technique Discussion Thread [Re: weetsie]
    #23501883 - 08/02/16 01:16 PM (7 years, 5 months ago)

Hah! I must of missed that bit  :canthearyou:


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Invisiblestareatclouds
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Re: Sterile Technique Discussion Thread [Re: Psykronik]
    #23501939 - 08/02/16 01:40 PM (7 years, 5 months ago)

1. Move diligently and with purpose.
2. Avoid putting things directly over an open plate/jar if possible.
3. 70% isopropyl alcohol and nitrile gloves.


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Re: Sterile Technique Discussion Thread [Re: stareatclouds]
    #23501969 - 08/02/16 01:49 PM (7 years, 5 months ago)

I really don't think skin is cleaner than anything you can put over top of it.  It's probably ejecting skin cells at a pretty insane rate just via normal activity.  A clean shirt would probably be best, but I always just do SAB work in whatever I'm wearing, and I don't shower or brush my teeth or put on anything beforehand other than nylon/nitrile gloves and a breathing/dust mask to diffuse my breath enough that it doesn't create currents.  None of those steps ever seem to translate into higher success rates, just a longer list of shit to do for no reason just to accomplish an inoculation.

The mask is only so I breathe normally without thinking too much about it, and the gloves are only because I'm not into spraying my bare hands with iso.


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Invisiblefilthyknees
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Re: Sterile Technique Discussion Thread [Re: Inocuole]
    #23502093 - 08/02/16 02:28 PM (7 years, 5 months ago)

Began using flow hood a month ago after using sab for a couple years.

I used to not shower, brush teeth or anything.
Now I try to view it as work. Not getting stoned helps.

Before doing any work with the hood I brush teeth, shower scrubbing everything, wet my hair, tie it back, use a fresh towel and like you stay nude. No shoes in the work area. No dirty work before sterile work (cutting grass or xyz)

Before I shower if I'm unloading a pc during the session I'll move it next to me in front of the flow sitting on a 5 gallon bucket and wipe the wing nuts and handles.

Once I'm showered and dried with a clean towel I put on gloves and a sergical mask, wipe down the working surface, separate what I'm working on from other stuff before opening the pc.

Right from the pc Into the flow I do not wipe bags or jars. If I'm doing g2g I'll use a fresh alc wipe (comes in 100ct) for the jar top, getting the rim once then clamping the wipe under the lid I unscrew the lid getting in where the lid covers.

Once I rotate once full rotation (left)under the lid I'll retighten and wipe the rest of the jar and set close to the hood. I'll soak my hands good with ISO and grabbing one bag set it to the right of the jar with the sealer to the right of the bag. Opening the bag angled towards the flow I'll keep fingers back as far as possible from the opening, loosen the master jar lid removing the top and with the left hand barely extend the rim over the bag enough to pour grain and no more, I'll twist and not shake the jar into the bag.

Grab the bag still angled towards the hood and keeping my hands back from the opening, close and turn to the right and seal.
Then I squeeze the bag just after sealing and listen for air leaks.

Wipe surface and repeat. I use a lot of alc wipes in a session.

For a2a I got lots of scalpels with fresh blades that if a pc is being run get sterilized.

I wipe the working surface, select the plate I'm starting on, wipe down the wrapped dish, unwrap it, and wipe it down again before opening. I don't cool the blade before transfers, no nessecary and a lot of extra hand movements when making 80 transfers like this morning.
Usually have a stack of 5-10+ receiving dishes to the left of the donor plate that are transfered to top down, restacking once transfered to more left close to the hood until I have 20 or more to wrap.
I'll wipe the parafilm box down and the cutting blades down with a fresh alc wipe, cut my strips, wrap, label and return to sleeve cover kept in front of flow hood.
If I have anything strange on the plate I'll keep it down stream of where I'm transferring from, and last out of any transfers to be done.


--------------------
But if you're in a hurry, and really got to go
If you're in a hurry, might have to find out slow
That it's one thing to try and another to fly
You get there quicker just a step at a time
It's one thing to bark, another to bite
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InvisibleMad Season
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Re: Sterile Technique Discussion Thread [Re: filthyknees]
    #23502219 - 08/02/16 03:08 PM (7 years, 5 months ago)

It really doesn't matter. Shirt and gloves or no shirt and no gloves. If you go over open media, or upwind of it, you're fucked either way lol. Wow SABs are fuckin awesome. Spray the walls and go to town! :rofl: that's literally my prep procedure lol.

I smoke like a chimney during SAB work so a face mask would end up on the floor anyways lol! Also cough a bunch too so I wouldn't worry too much about sab air currents from breath :hehehe: unless you're like an extremely obnoxious breather. I also got my laptop on top playing a playlist of movies :lol: I mean if you're in there for a few hours, gotta make some fun out of it. Compared to some of you guys, I'm such a slob :lol:


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InvisibleInocuole
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Re: Sterile Technique Discussion Thread [Re: Mad Season]
    #23502224 - 08/02/16 03:09 PM (7 years, 5 months ago)

Most such things are only for peace of mind and don't really help. :shrug:


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OfflinePsilocyBen17
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Re: Sterile Technique Discussion Thread [Re: Inocuole]
    #23502247 - 08/02/16 03:15 PM (7 years, 5 months ago)

Quote:

Inocuole said:
I really don't think skin is cleaner than anything you can put over top of it.  It's probably ejecting skin cells at a pretty insane rate just via normal activity.  A clean shirt would probably be best, but I always just do SAB work in whatever I'm wearing, and I don't shower or brush my teeth or put on anything beforehand other than nylon/nitrile gloves and a breathing/dust mask to diffuse my breath enough that it doesn't create currents.  None of those steps ever seem to translate into higher success rates, just a longer list of shit to do for no reason just to accomplish an inoculation.

The mask is only so I breathe normally without thinking too much about it, and the gloves are only because I'm not into spraying my bare hands with iso.




This is really helpful. Thanks innoc. Good point about the clean shirt business.


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OfflinePsilocyBen17
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Re: Sterile Technique Discussion Thread [Re: Mad Season]
    #23502261 - 08/02/16 03:20 PM (7 years, 5 months ago)

Quote:

Mad Season said:
It really doesn't matter. Shirt and gloves or no shirt and no gloves. If you go over open media, or upwind of it, you're fucked either way lol. Wow SABs are fuckin awesome. Spray the walls and go to town! :rofl: that's literally my prep procedure lol.

I smoke like a chimney during SAB work so a face mask would end up on the floor anyways lol! Also cough a bunch too so I wouldn't worry too much about sab air currents from breath :hehehe: unless you're like an extremely obnoxious breather. I also got my laptop on top playing a playlist of movies :lol: I mean if you're in there for a few hours, gotta make some fun out of it. Compared to some of you guys, I'm such a slob :lol:




I was such an idiot back when I was using a SAB....I really wish I had have understood what I was doing better and worked on refining my technique....

So you just spray the walls with water and stay off the floor as much as possible? Do you load it up with jars before you spray? You dont find spraying gets your filters wet?

You would just put the jars and tools on a table and then place the SAB over top of it right?


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Re: Sterile Technique Discussion Thread [Re: PsilocyBen17]
    #23502276 - 08/02/16 03:26 PM (7 years, 5 months ago)

If spraying gets your filters wet in your mind, then spray the box before you put the jars in them. :lol:  Wet jars are slippery and unwieldy.  Only the box and floor need to be wet.


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Re: Sterile Technique Discussion Thread [Re: PsilocyBen17]
    #23502283 - 08/02/16 03:30 PM (7 years, 5 months ago)

Filters getting wet is not a Big deal. Filters staying wet allowing contams to grow in/through them is a problem.


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Re: Sterile Technique Discussion Thread [Re: Pastywhyte]
    #23502397 - 08/02/16 04:01 PM (7 years, 5 months ago)

A couple times iv had a drop of water go from the roof of my Sab right onto a plate. I need to spray less water I guess


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Re: Sterile Technique Discussion Thread [Re: Pastywhyte]
    #23502408 - 08/02/16 04:04 PM (7 years, 5 months ago)

I've found the more I post  sarcastic comments on forums,
inbetween not showering and sterile work,
The better my grows get.
Can anyone confirm this or is it just me?
:blazed:


--------------------
But if you're in a hurry, and really got to go
If you're in a hurry, might have to find out slow
That it's one thing to try and another to fly
You get there quicker just a step at a time
It's one thing to bark, another to bite
The show ain't over till you pack up at night


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Re: Sterile Technique Discussion Thread [Re: filthyknees]
    #23502449 - 08/02/16 04:15 PM (7 years, 5 months ago)

Quote:

filthyknees said:
I've found the more I post  sarcastic comments on forums,
inbetween not showering and sterile work,
The better my grows get.
Can anyone confirm this or is it just me?
:blazed:




Nah dude that's like, THE method.  All the pros do it.


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Re: Sterile Technique Discussion Thread [Re: Inocuole]
    #23502773 - 08/02/16 05:57 PM (7 years, 5 months ago)

Used to have big problems with sterility as I have no access to a room that even remotely suits a lab room.
My kitchen is the only place that is half suitable, and that connects all the other rooms to it and is very untidy and dirty most of the time. Have since refined technique and now use a tyvek suit/gloves/face mask together with lots of 70 alcohol and lots of wiping both inside and outside of the SAB.
ive never bothered with showering and all tthat jazz. I did however have a problem with all my last batch of agar plates but I think that's due to the fact my PC ran out of water at the 12 minute mark and I had to run and turn it off :frown:
In summary, I'd say if you guys use a room that you are able to keep relatively clean and tidy when doing work, ditch the whole shower, wash hair etc etc and just get a tyvek suit. Breaking bad ftw


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OfflinePsilocyBen17
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Re: Sterile Technique Discussion Thread [Re: deagle5]
    #23502840 - 08/02/16 06:21 PM (7 years, 5 months ago)

No way dude. I put on a tyvek suit once and my balls got so crushed that they were mad at me for weeks...not to mention you sweat like a motherfucker, I'd be done after being in one of those things for 10 minutes.

what im getting from this conversation is that if you work in a SAB or infront of a bench than showering and teethbrushing are just for bonus feel good points.


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InvisibleInocuole
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Re: Sterile Technique Discussion Thread [Re: PsilocyBen17]
    #23502859 - 08/02/16 06:25 PM (7 years, 5 months ago)

Basically.  You'd have to be real nasty for a tyvek suit to be the thing that increases your success rate.


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                            :rainbowdrink: Tea doesn't work?                            AMU  (Q & A)                  Grain prep for Intergalactic Space Oats :pes:     

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OfflineKKVguitarist
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Ultrasonic... would this work? [Re: PsilocyBen17]
    #24118492 - 02/25/17 10:17 AM (6 years, 10 months ago)

In order to sterilize a fruiting chamber before use( while its empty) with all needed supplies inside, and im talking about a martha but any could work, what if you put straight rubbing alcohol inside an ultrasonic humidifier and let it run in a sealed environment so every nook and cranny got sterilized? I dont see why this wouldnt work then allow to dry. Could be a good step just to ensure you start off on a good note


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