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OfflineSMmack
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Consistent contam. Critique my technique. * 1
    #28688794 - 03/06/24 02:04 PM (10 months, 6 days ago)

So I haven’t’ been able to grow anything over the past year. My dirty UB tek in 2022 worked better than anything I’ve tried since starting agar. I’ve had a lot of help from you all, and have listened to it all, but am still getting  what I think is a ridiculously disproportionate amount of contam for the methods I’m using, and I can’t figure out why. Early last year I was having a nightmare with a fungicola strain almost identical to tomentose mycelium which got entangled in my strains, this lead to growing a lot of fungicola and not a lot shrooms. Around summer I finally got mushroom mycelium but had something growing under it making it stall. I think it may have been more fungicola but who knows. These all had various contam but something similar when sending to spawn… Trich. Over winter I had changed most of what I was doing and was getting mostly just trich and a bit of that super dense white shit that goes trichy as soon as you open it, not much sour rot, cobweb or black anymore. So in regards to consistency, I’ve made some progress, but now trich seems more prevelant than ever. Before I was getting some plates to come out clean, now almost every plate is contaminated. You guys keep asking about different parts of my technique so I thought I’d make an overview post to show everything that I do, start to finish. If I could get any feedback or critique on it that would be so helpful. This is going to be long but with lots of photos, so I’ll subdivide it into headings and try and keep it as concise as possible, with all the info I’m usually asked.

The headings:
Feel free to skip through any that you don’t feel are relevant.
- Recent examples
- Agar Prep
- SAB Prep
- SAB Technique
- Results of SAB transfers

First some of my recent works:
These are my old cups, stored upside down and not so wet as to cause problems, I test everything first so all contam has been introduced since introduction to the SAB. None of these transferred to be clean, but I’m not convinced that’s not from introduction of contam rather than these being covered in trich.

Someone noted they were getting trich in the lid gap when the condensation dripped from agar to lid, which would harbour nutrients and then make its way up into the edge of the agar, so I recently started storing the cups right side up…

20/20 plates were like this, and the same on batch two the next week, although a bit better.



So let’s look through my methods.



Agar prep:
1L water, 20g LMEA, 20g agar, gel colouring. Mix under heat until start of boil.
10ml in each puck or plate. The extra goes in jars straight into the freezer for future use.

Stack in PC. I modified the lid, turning 12psi to 15psi. DO NOT DO THIS IF YOU DONT KNOW WHAT YOUR DOING. I still pc for the time of a 12psi for sanity reasons, this just gives me extra killability but is probaly not needed.

Vent once lock pops up for 15 mins.
Pc for 45mins-1hr.
Take out 30 mins after lock drops, wipe off liquid on lids and spread out to cool and reduce condensation.
Store in cupboard for a few days until confirmed no growths, add 2 more layers of micropore tape to air holes if plastic and move to SAB.



This batch was fairly average my no pour pots had a 100% success rate, but quite a lot of condensation asside from the ones on the top of the pile. Two of my glass Petri dishes have contam and the bottom two on each pile came out empty.
Plates moved to the top have no agar anymore.

What I think happened to introduce contam to those two petris was that there’s nothing sealing the petri’s so when I take them out of the PC to reduce condenstation they have a chance to break the liquid seal and open slightly, this is why I moved to no-pour pots, contam rates can be reduced by leaving until cold and moving the whole stack directly to storage cupboard, but then condensation is a lot higher. The latter problem is that the the bottom ones boil over after taking most of the heat, thing is there’s now agar nutrient that has come out of the lid, so its contam prone at the sides. The main thing I don’t like about them is that if they can contam while handling out of PC then they’re likely to contam while handling after inoculation, I wrap 3 times with cling film to help prevent it, but that time in between I’m convinced is where my fungicola saga originated early last year.



SAB Prep:
100qt box, on side with 6” holes cut out.
Clinell wipe SAB, spray 10% bleach and wipe SAB, 70% iso spray and wipe SAB (because a little mustard gas never hurt anyone). Transfer plates from cupboard to SAB, anything trichy stays out until all the clean stuff is already done. Lid on. Spray whole SAB with 70% iso, heavy books to prevent movement & leave for minimum of 20 minutes. From here there is no more spraying of iso except on my gloves every now and then.
Handy tip, I whiteboard marker on the lid what I’m planning to transfer, and wipe off after doing each one, this prevents the problem of forgetting to make some transfers or making too many of one kind.



Doff clothes, Don mask and gloves, then stand over the SAB looking down through the top.


SAB Technique
This week I’m doing my usual technique but trying a bit of everything in terms of storage, troubleshooting. I’m going back to my old glass Petri dishes, storing some no-pour pots upside down and others right side up. I’m using the same pots as earlier just more developed now, and I’ve done some isolating of  some new but very contaminated plates, just to see if that’s any more reliable.

Let’s get to work.

Transfer donors Old samples of spores T0 from last month. Labelled A-F. I transferred to:
- 1 upright pot
- 1 upside down pot
- 1 Petri dish upright
As you can see there’s shit in the lids, whether that’s just the same amount of contam as the upright pots, but not on the agar because they were all stored upside down, I can’t tell you.



Transferring
The video shows my technique during a full transfer, a little bit more clunky because of the camera. Usually if I’m doing multiple transfers to the same plate or from the same donor area, I will only flame between plates, so doing two transfers per flame instead of one. Any time I’m going to a different location or doing more than two transfers I’ll always flame, but if I start a transfer and cut two wedges out, the second wedge is pre cut and I get very little additional contam in this case when compared to the extra time of flaming while the lid is off. I probaly should have flamed between the first two plates but I was running low on time and I did two wedges on the first cut out of habit. The contam is affecting both pots equally when compared to flaming between each pots.




This video shows how I flame the blade. I dip it in a jar of 70% after transferring to get the agar off, then take out and torch. The 70% sets alight the handle while the blade gets red hot, transfer back to SAB before letting cool down.



This video shows how I put the lid down if I need a better view than just cracking the lid open. I’d rather crack the lid, but in the event I need to transfer away from a contaminant and accuracy is key, I’ll flip the lid over, keep working at the back of the SAB, and crack the target plates.



These are the more recent contaminated samples since I started storing right side up, A-F T1 and also a pin clone from an old BRF puck. Let’s call them the dirty plates. I transferred to one Petri dish each. They don’t look great, but I’ve managed to clean up worse than this before.
\



Incubation
All transfers go in a box in the boiler cupboard, it allows some gas exchange but is not subject to daily movements. It also maintains a nice 18-21˚C, so a little cold compared to my heat mat cupboard, but I’ve had some problems with that making jars too wet in the past. So I tend to stick to the storage box.






Results
First my transfers from the older clean looking samples;
Condensation pooling in all the tubs .


Right side up tubs: A B C D E F
There’s nothing growing in the lid gaps, as we’ll see later, and the contam doesn’t seem to show any correlation to where the micropore tape holes are, if it was coming through there I would  assume there to be more growth just below the hole.


Upside down tubs: A C
D
E , F

There’s a lot more to see here. On A there are a few tiny dots and some light powdery stuff in the lid gap but nothing obvious yet, A seems to appear the cleanest but usually catches up to the others over time. D is where we really start to see stuff without having to look closely, there’s contam growing in the water on the surface of the lid, and there’s contam growing in the water in the gap between lid and pot, this is condensation, but if it has come from the agar it would have nutrients in. It is plausible that contam may be able to follow the trail of rich condensation to the agar, however the spread of colonies dosen’t seem to fit such a narrative, I’d expect a growth starting from the edge if that were the case. The side photo of F is probaly one of the easier photos to see what’s going on in the lid gap.


Petri Dishes right side up:
A C D E F .
A looks clean. CDE are trichy, F trich again and some bacc physically transferred.



After I took my clean plates out, put my other plates in and transferred from those dirty trich plates:

Clone of a pin:
Stored Right side up. Stored Upside down. In a Petri Dish. Another plate other side same plate

A spot of trich, and some ?clear slime to the side of the first one (an outlier I think). And a clear Petri dish.


Now trying to isolate the recent but very trichy plates:
A C D E F

C is a plate with agar boiled away, very little agar left and very little myc growth with it. That said, not much trich even though it has no problem growing on a super thin layer of agar. E is another really thin layer a few spots of trich, but nothing is going to grow on this. F one spot of trich, one random spot of bacc in the middle.

So my pots of ultra sporulating trich are less contam prone than my other clean looking plates, so I wasn’t expecting that, and now I’m even more confused than when I started.


This is still early days, I’m expecting a couple more plates to go bad in the coming week, but I think we can see a few trends in place. Given the seemingly equal spread of trich, I’d say its airborne entry, however given that my trichy AF transfers ended up cleaner than my clean transfers that might suggest the trich contam isn’t getting in via SAB, as I’d expect the contam to get worse as work time increased and when I introduced sporulating trich to SAB. My large surface area Petri dishes also seem to get less contam, which might suggest it’s getting through the lid gaps of the pots, and somewhat through the petri’s aswell.

Here’s a spawn jar from A
Given the location of trich is not intertwined with the healthy myc, I’m convinced the original donor pots are clean enough, and somewhere along the line there’s shit getting in. The problem is I don’t know where.

Maybe the trich is getting through the lids of the pots and spreading to others nearby, that doesn’t explain the equal surface spread or that the wrapped petris are getting it. Maybe it’s coming through the micropore, again doesn’t explain the spread or petri’s. Maybe my PC is bad, dosen’t explain why all of the used plates passed QC. So I think that just leaves introduction of contam via SAB technique.


Do you see what I mean about consistent trich? Do you see any patterns? Where do you think this is coming from?
IDK, What would you do if faced with these results?

Edited by SMmack (03/06/24 02:09 PM)

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InvisibleBaba Yaga
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Re: Consistent contam. Critique my technique. [Re: SMmack] * 1
    #28688849 - 03/06/24 03:09 PM (10 months, 6 days ago)

Hey mate, good to see you have done the detailed write up on your sterile work. Ironically I didn't have the time to read it all and just looked at the vids as they are the best thing to give insight. I know what you are doing from our previous discussion anyway.

IMO your movements are too fast and uncoordinated in general and when you going out and coming back in. Your hands are going back and forth more than they need to.

The way you open the donor plate is not good IMO, lots of rubbing/friction of the gloves on the rim of the cup and too much glove too close to the cup. I would use the donor upside down instead or build something to hold it while cutting the wedge so you can lift off the lid. Same for when you are opening the receiving plates, it does look like the fingers which are not touching the lid are touching rubbing against the rim of the cup. Only touch the lid with two finger tips and keep the rest out of the way/away from the cup.

This is how I do it, it's not filmed in a SAB to better show the movements.




I think you need a bigger SAB or do not have as much stuff in there. The petri dishes stacked in the back of the SAB is valuable space wasted as it pushes your work area closer to the front and opening and it seems you are already working quite close to the front. You want to work as far back as possible and the plates are in the way. Stack them to the side if you can or leave them out. It's better to split the session instead of overfilling the SAB. You only have to set up the SAB once and then take stuff out that is done and put new stuff in.

Your gloves look dry. Also wondering if they are powdered inside. Have you tried working other gloves or without?

Well, it's mostly movement and coordination as far as I can see.

Edited by Baba Yaga (03/06/24 03:38 PM)

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InvisibleLadysKnight
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Re: Consistent contam. Critique my technique. [Re: SMmack] * 2
    #28688861 - 03/06/24 03:19 PM (10 months, 6 days ago)

I read pretty far and several things were getting off track, like pulling no-pours out of the PC while hot, then leaving them on the counter til cool, and you are cooking them too long and boiling them out. So instead of addressing individual issues, I see you have some glass plates. You should try this basic, standard tek and report back:
https://www.shroomery.org/forums/showflat.php/Number/28006384

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InvisibleDaeda1us
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Re: Consistent contam. Critique my technique. [Re: LadysKnight] * 1
    #28688863 - 03/06/24 03:21 PM (10 months, 6 days ago)

very detailed post, i may have missed a detail or two. trying to pinpoint where your fatal flaw is, still unsure. couple things i noticed:
  • be careful of your movements in the SAB. as a general rule, nothing non-sterile above sterile. this includes your hands. plan your placement of objects and movements in advance.
  • you're not wrapping your pasty plates in foil while pcing? 
  • pics seem to indicate you're doing no pour glass petris? is that even a thing?


seems to me you have an error with your PC process or your sterile technique.  i use regular vented 100x15mm petris. i don't wrap the plates after i pour. i usually use the plates a week after pouring, then wrap them.

for pasty plates, i would keep the foil on until you're loading the SAB.


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InvisibleDaeda1us
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Re: Consistent contam. Critique my technique. [Re: LadysKnight] * 1
    #28688882 - 03/06/24 03:30 PM (10 months, 6 days ago)

thank you for that link, will for sure be using it. why did i spend that money on glass plates instead of pp5?

oh well, live and learn. will get some use, then maybe i'll load them with orange agar & use as clay pigeons.


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InvisibleBaba Yaga
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Re: Consistent contam. Critique my technique. [Re: LadysKnight] * 1
    #28688899 - 03/06/24 03:36 PM (10 months, 6 days ago)

Quote:

LadysKnight said:
I read pretty far and several things were getting off track, like pulling no-pours out of the PC while hot, then leaving them on the counter til cool, and you are cooking them too long and boiling them out. So instead of addressing individual issues, I see you have some glass plates. You should try this basic, standard tek and report back:
https://www.shroomery.org/forums/showflat.php/Number/28006384




I agree that the PC time is too long but taking them out hot as he does is fine. I follow mostly Josex's write up on those condiments cups and always take them out hot. About 20 min after the pressure indicator drops and then stack them to reduce condensation on the lids. Not a problem.

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InvisibleLadysKnight
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Re: Consistent contam. Critique my technique. [Re: Baba Yaga] * 2
    #28688931 - 03/06/24 03:59 PM (10 months, 6 days ago)

Quote:

Baba Yaga said:
Quote:

LadysKnight said:
I read pretty far and several things were getting off track, like pulling no-pours out of the PC while hot, then leaving them on the counter til cool, and you are cooking them too long and boiling them out. So instead of addressing individual issues, I see you have some glass plates. You should try this basic, standard tek and report back:
https://www.shroomery.org/forums/showflat.php/Number/28006384




I agree that the PC time is too long but taking them out hot as he does is fine. I follow mostly Josex's write up on those condiments cups and always take them out hot. About 20 min after the pressure indicator drops and then stack them to reduce condensation on the lids. Not a problem.



Yeah it seems he's using the condiment cups and glass plates and mixing teks. With glass, I leave them in the PC til it's cold so the agar is stiff then they go straight to the sab to be used. Never do they hang out on the counter or anywhere in the open before use/wrapping.

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InvisibleBaba Yaga
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Re: Consistent contam. Critique my technique. [Re: LadysKnight] * 2
    #28688987 - 03/06/24 04:42 PM (10 months, 6 days ago)

Yeah, wouldn't do that with glass petris either

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OfflineSMmack
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Re: Consistent contam. Critique my technique. [Re: LadysKnight]
    #28690316 - 03/07/24 04:23 PM (10 months, 5 days ago)

Quote:

LadysKnight said:
Quote:

Baba Yaga said:
Quote:

LadysKnight said:
I read pretty far and several things were getting off track, like pulling no-pours out of the PC while hot, then leaving them on the counter til cool, and you are cooking them too long and boiling them out. So instead of addressing individual issues, I see you have some glass plates. You should try this basic, standard tek and report back:
https://www.shroomery.org/forums/showflat.php/Number/28006384




I agree that the PC time is too long but taking them out hot as he does is fine. I follow mostly Josex's write up on those condiments cups and always take them out hot. About 20 min after the pressure indicator drops and then stack them to reduce condensation on the lids. Not a problem.



Yeah it seems he's using the condiment cups and glass plates and mixing teks. With glass, I leave them in the PC til it's cold so the agar is stiff then they go straight to the sab to be used. Never do they hang out on the counter or anywhere in the open before use/wrapping.




Yeah normally I do one or the other. In this batch I wanted to troubleshoot so did a bit of everything. Glass petris I’d normally leave in the pc and not open until use. The sealed pp5 ones I take out whenever. Taking the glass out with the pp5 was obviously what gave me contam in 2 of them, but that’s why I left them out for a week to check I only used clean ones.

I’m doing my next transfers this weekend, so I’ll ill follow with petris and report next week. Or maybe just do them both but separate batches, I only have 20 petris…

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OfflineSMmack
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Re: Consistent contam. Critique my technique. [Re: Daeda1us]
    #28690331 - 03/07/24 04:31 PM (10 months, 5 days ago)

Quote:

Daeda1us said:
very detailed post, i may have missed a detail or two. trying to pinpoint where your fatal flaw is, still unsure. couple things i noticed:
  • be careful of your movements in the SAB. as a general rule, nothing non-sterile above sterile. this includes your hands. plan your placement of objects and movements in advance.
  • you're not wrapping your pasty plates in foil while pcing? 
  • pics seem to indicate you're doing no pour glass petris? is that even a thing?


seems to me you have an error with your PC process or your sterile technique.  i use regular vented 100x15mm petris. i don't wrap the plates after i pour. i usually use the plates a week after pouring, then wrap them.

for pasty plates, i would keep the foil on until you're loading the SAB.




Foil on for the cook generally. Sometimes I forget but it only seems to prevent accidentals.

For the amount of contam I get from a few seconds of SAB use I wouldn’t trust pouring. There’s no reason you can’t no-pour them, people don’t typically do it because the plastic ones don’t hold up in PC, if your autoclaving glass ones for use anyway you might aswell put the agar in prior and skip a step, just be careful you don’t get paranoid and boil them over too long.

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OfflineSMmack
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Re: Consistent contam. Critique my technique. [Re: Baba Yaga]
    #28690365 - 03/07/24 04:45 PM (10 months, 5 days ago)

Quote:

Baba Yaga said:
Hey mate, good to see you have done the detailed write up on your sterile work. Ironically I didn't have the time to read it all and just looked at the vids as they are the best thing to give insight. I know what you are doing from our previous discussion anyway.

IMO your movements are too fast and uncoordinated in general and when you going out and coming back in. Your hands are going back and forth more than they need to.

The way you open the donor plate is not good IMO, lots of rubbing/friction of the gloves on the rim of the cup and too much glove too close to the cup. I would use the donor upside down instead or build something to hold it while cutting the wedge so you can lift off the lid. Same for when you are opening the receiving plates, it does look like the fingers which are not touching the lid are touching rubbing against the rim of the cup. Only touch the lid with two finger tips and keep the rest out of the way/away from the cup.

This is how I do it, it's not filmed in a SAB to better show the movements.




I think you need a bigger SAB or do not have as much stuff in there. The petri dishes stacked in the back of the SAB is valuable space wasted as it pushes your work area closer to the front and opening and it seems you are already working quite close to the front. You want to work as far back as possible and the plates are in the way. Stack them to the side if you can or leave them out. It's better to split the session instead of overfilling the SAB. You only have to set up the SAB once and then take stuff out that is done and put new stuff in.

Your gloves look dry. Also wondering if they are powdered inside. Have you tried working other gloves or without?

Well, it's mostly movement and coordination as far as I can see.





This is a long narrow SAB, there’s a lot more room behind the camera, my arms are fully in there. I will use that space more effectively though. Plates on the side next time.

If you didn’t read the post I don’t blame you. The TLDR is that it’s definitely introduced during transfer and not my PC prep, I just wasn’t able to pinpoint what the catastrophic faliure point was.

I’ll get some empty pots and practice to your gif before next transfers this weekend. After working in a flow hood I’ve developed a big meaty grip. You can spit in those things and not contaminate anything.



So for the pastywhites, store right side up, but flip upside down when transferring, then back to right way up?

How is the petri dish opening also? Obviously a lot easier and smoother to open as they don’t seal. I did report a significantly lower contam to surface area ratio when compared to the pasty’s but most of them still got around 1-3 trich spots on the plate. I also need to get some of the old cling film that actually sticks, this new eco friendly stuff is driving me nuts.

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InvisibleLadysKnight
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Re: Consistent contam. Critique my technique. [Re: SMmack] * 1
    #28690566 - 03/07/24 06:50 PM (10 months, 5 days ago)

You said when you use glass plates you leave them in the PC until you're ready to use them.

I would move them from the PC to sab when cool. Then use and wrap.

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Re: Consistent contam. Critique my technique. [Re: SMmack]
    #28691149 - 03/08/24 06:43 AM (10 months, 4 days ago)

I'm also currently reviewing my sab work asm before getting back. Nice detailed write up for debug(for you & me aswell).

Just to add up,

Your scalpel flaming is bueno, I also do that alcohol dip, except mine is outside of the sab (quick dip then flame) for less clutter.

Though it's a good thing you had yours(iso dip) inside the sab for debugging in the video. I had to use a water bottle for this.

Seems your working on a dry sab, I also - sometimes do. Being Dry(or even not) I'd avoid vibrating the sab as much as seen @ 3:25(goin in) specially @ 2:35 (snapping lids of receiving cups). I could imagine that dry sab floor bouncing those spores back.

I stack my receiving cups like in baba's gif (to keep further away from the floor) for transfers.
I don't immediately snap each lids after each xfer, just doing it after I made all xfers near the end of the sab work(lifting each cup to snap the lid).
I also wrap them in my part(before PC and after working with them) since some of my lids are a bit loose.


Edited by pacmanbreed (03/08/24 07:23 AM)

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OfflineDERRAYLD
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Re: Consistent contam. Critique my technique. [Re: pacmanbreed] * 1
    #28691162 - 03/08/24 07:10 AM (10 months, 4 days ago)

Sorry but ill admit that was too much for me to read so I just looked at what I know to be true and the faults in your photos.

You must wrap petris in foil when pcing them or they will suck in contamination when they cool inside the pc.
The same applies to deli cups from my experience.

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Re: Consistent contam. Critique my technique. [Re: DERRAYLD] * 1
    #28691425 - 03/08/24 11:47 AM (10 months, 4 days ago)

Admittedly I did not read this whole thread, but I have a few suggestions/comments. (Note that I’m VERY new to mushroom cultivation, but I’m having >90% success with agar transfers in a SAB so I think I have some ground to stand on.)

* Maybe I missed it, but are you misting the interior of your SAB with water/soap solution and letting that settle out before putting anything in there?

* I use screw-top agar containers, but I think the process is very similar overall. I’ve been aiming to keep my gloves visibly wet with 70% ISO whenever I’m screwing/unscrewing the tops. The I leave the tops loosely in place so I can minimize the amount of time the plates are exposed to air.

* I would aim to work further back in your SAB. You’re leaving your donor plate wide open in the front area of the SAB where ambient dust is most likely make it’s way into the arm holes - especially when you’re being your right hand in and out of the SAB to flame the blade, etc.

* I noticed you’re transferring from a plate that looks fully colonized. It’s not surprising to me that you’re grabbing mycelium AND contamination when both have had plenty of time to cover the surface of the agar. Instead, I’d recommend transferring much earlier. Grab the very leading edge of the fastest/furthest growing mycelium where it has “out run” the contamination. I try to grab the tiniest little bit from the edge of the white, or even a little in front of the white visible part (most of the bits I transfer are about half white and half clear, and about the size of a grain of rice - pretty much as small as I can go).

Here’s what I do:

1)  Loosen the lid of your donor plate using ISO soaked (but not dripping…) gloves. Leave the lid in place. Same for your receiving plate.

2) Flame your blade.

3) Barely crack the lid of your receiving plate, dunk the blade into the agar to cool it, then carefully lower the lid (without clicking it shut).

4) Carefully lift off the lid of the donor plate and place it on your work sanitized work surface (do not flip it - keep the clean side facing down so contamination can’t land on it). Cut your sliver of agar, then, using your left hand, carefully place the lid back on the donor plate (make sure you place the lid straight down - try not to rub the edges of the lid where it has contacted your work surface against the lip of the container).

5) Barely crack open the receiving plate and deposit the sliver of agar in the middle without ever holding passing your hands above the clean agar surface.

Overall, I think your problem is the way you’re leaving your donor plate wide open right near the arm holes. I think you’ll have better luck if you do everything g possible to minimize the time that plates are open and exposed to the elements.

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Re: Consistent contam. Critique my technique. [Re: SMmack] * 1
    #28691506 - 03/08/24 12:39 PM (10 months, 4 days ago)

It is my understanding that alcohol does not kill mold spores. you can sterilize your scalpel by quickly torching the last 3 or 4 inches of the handle before getting the blade red hot. this will kill anything on the surface of the scalpel more effectively than burning off an iso-soaked handle.

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Re: Consistent contam. Critique my technique. [Re: pacmanbreed] * 1
    #28691712 - 03/08/24 03:43 PM (10 months, 4 days ago)

Quote:

pacmanbreed said:
I'm also currently reviewing my sab work asm before getting back. Nice detailed write up for debug(for you & me aswell).

Just to add up,

Your scalpel flaming is bueno, I also do that alcohol dip, except mine is outside of the sab (quick dip then flame) for less clutter.

Though it's a good thing you had yours(iso dip) inside the sab for debugging in the video. I had to use a water bottle for this.

Seems your working on a dry sab, I also - sometimes do. Being Dry(or even not) I'd avoid vibrating the sab as much as seen @ 3:25(goin in) specially @ 2:35 (snapping lids of receiving cups). I could imagine that dry sab floor bouncing those spores back.

I stack my receiving cups like in baba's gif (to keep further away from the floor) for transfers.
I don't immediately snap each lids after each xfer, just doing it after I made all xfers near the end of the sab work(lifting each cup to snap the lid).




I never thought to use water to detect vibrations.
I’m also going to add a filter disk cushion to the bottom of the iso dip because metal against glass vibes a lot and also bends the blade.

So loosen the lids in the air, stack pastywhite’s, wait 5, and work down the tower?

I wipe bleach, wipe iso, then spray iso before use. From there it stays dry.

By wet SAB do you mean the soapy water tek? I tried that but with the middle of the SAB being the lowest point it kept dripping into my work. Would a wet soapy paper towel on the floor and soapy spray on the walls would be better. Bleach and iso to clean, then a quick soap setup before adding things inside?

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Re: Consistent contam. Critique my technique. [Re: SMmack] * 1
    #28691777 - 03/08/24 04:49 PM (10 months, 4 days ago)

ISO, bleach, spraying, wiping, you need to understand that is done to give YOU peace of mind. it only does a little to reduce contamination rates. The #1 thing to do with a STILL AIR BOX is to keep the air still. IMHO, you are using the STILL AIR BOX wrong. we do all of our work in the bottom of the STILL AIR BOX so let's focus on that. You have a rack elevating your work, that's good. unfortunately, it's setting directly on a side of your box, that's bad. Every vibration will transmit throughout the entire box, stirring up the air and dislodging contaminants. GOOD NEWS, the solution is simple! Get rid of the lid you have cut armholes into. Lay out a towel larger than the top of your SAB, this IS NOT used to "trap contaminates", its purpose is to muffle any vibrations and to seal the edges from any air drafts (some people use weather stripping instead), no need to wet it, let gravity do its job. Almost done! Flip the box so the open top sets on the towel, cut new, large arm holes, put those books on a bookshelf, and you're done! A box full of still air, gravity doing all the work of cleaning the air inside, sweet.  (oh yeah, armhole covers are good to have, almost forgot that)

Edited by normalperson (03/08/24 04:52 PM)

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Re: Consistent contam. Critique my technique. [Re: SMmack]
    #28691892 - 03/08/24 06:21 PM (10 months, 4 days ago)

:whathesaid: nice minimal sab setup.

Good summarized explanation, I'll add that to what TC fellows & others already mentioned.

  • Baba - gentle hand movements
  • DERRAYLD - wrap cups/glass petris (before PC and after) - to further minimize contamination factors.(I cling wrap mine most of the time)
  • Flame the handle for a bit - from my exp aswell


I'd flip that sab upside down and toss the lid as per recommended.(The lower the arm holes and the more elevated the work area the better)

But since we both have the same wrong arm hole placement, and if cant be replaced/repaired as of the moment, avoid touching/vibrating sab walls can help.


Quote:

I never thought to use water to detect vibrations.
I’m also going to add a filter disk cushion to the bottom of the iso dip because metal against glass vibes a lot and also bends the blade.



just stumble upon - I also never thought of it aswell. From my limited understanding - gas acts like fluid, the sab floor is ladden with spore/contamination. If you can iso dip & blade swirl comfortably outside sab - the better, minimizing the factor(vibrating the sab walls), Its one of the multiple reasons I do it outside aside from ease.

just be careful not to let the hand soak iso catch fire, I have experienced this quite often in the past - doing it in quick succession, being carried away by the work flow - letting the iso from the blade creep thru my hands while heating it! (as I love the poof sound of iso ladden blade on fire) - one of the multiple reasons mentioned :grin:.

Letting it drip for a second before Flaming it an angle below the hand level helps.


Quote:

So loosen the lids in the air, stack pastywhite’s, wait 5, and work down the tower?



Yes just like how petri plates are treated. Except if your comfortable enough with your hand movements, I do lift mine one by one from the stack - then restack them from up to bottom, noccing them sideways the way josex does(make use of gravity), this is all done at the furthest back of the sab as far as my hand can reach.


Quote:

By wet SAB do you mean the soapy water tek? I tried that but with the middle of the SAB being the lowest point it kept dripping into my work. Would a wet soapy paper towel on the floor and soapy spray on the walls would be better. Bleach and iso to clean, then a quick soap setup before adding things inside?



Yes - soapy water Tek, that's the way I do mine from before, expect I don't wipe it with iso quite often before setup, I directly mist it soapy water(Abit of bleach) but not soaking wet.
I've also experience that dripping(hence I sometimes work with dry sab when doing small amounts). As per soapy Tek - just spray & wipe the tote lid dry before closing the sab. And avoid vibrating it as much, during work.

Hope you can polish yours as-well, (goodluck).

Edited by pacmanbreed (03/08/24 07:53 PM)

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