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OfflinePhony Phone

Registered: 09/19/20
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SAB microparticles flying around, problem
    #27006378 - 10/27/20 01:35 PM (3 years, 3 months ago)

TL;DR: SAB micropatricles still flying inside it after precautions being taken, agar plates are too wiggly, need advice on how to handle.

============================================

After setting up the sab and spritzing it with soapwater, both the box and the towel, and also spraying the air with water, I decided to check how the conditions inside the box were.

So i took a flashlight and shone it inside the SAB. What i realized was that microparticles were STILL flying around, even after all precautions were taken. Now the box is supposed to be still air and the drafts in the room had all stopped hours ago so i have 0 clue as to why this phenomenon occurs.

I really appreciate some help on the matter.

Edit: I also wonder how you people open your petri sleeves inside the sab without the top part of the dishes wiggling around and being in danger of contamination?
And how do you grab the bottommost dish when you use say a cookie rack that gives no space for fingers to grab below it? Petris seem to be so wiggly and easy to open up by accident thus ruining the whole dish, combined with the fact of observing the microparticles flying around, the chances seem to lower dramatically.

Thanks for any information


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Offlinemad_dingo
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Registered: 08/19/20
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Re: SAB microparticles flying around, problem [Re: Phony Phone]
    #27006444 - 10/27/20 02:12 PM (3 years, 3 months ago)

Quote:

Phony Phone said:

And how do you grab the bottommost dish when you use say a cookie rack that gives no space for fingers to grab below it? Petris seem to be so wiggly and easy to open up by accident thus ruining the whole dish, combined with the fact of observing the microparticles flying around, the chances seem to lower dramatically.

Thanks for any information




I place my petri dishes on metal cookie cutters. That allows me to place my fingers on the bottom without smearing them all over the lid.

But i think with those petri dishes you just have to live with the probability the sooner or later contams will infiltrate from the side, especially if you have lots of condensation.


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Invisiblecoversall
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Re: SAB microparticles flying around, problem [Re: mad_dingo]
    #27006449 - 10/27/20 02:15 PM (3 years, 3 months ago)

My SAB has got so much easier to work in since getting a wire rack to go in the bottom. Maybe I'm not fully understanding what you say about the rack. But I slide my petris over to the edge of the rack then I can pick them up easily.

How long had your SAB sat before you checked it with the torch?

Have you been have contam issues?


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OfflinePhony Phone

Registered: 09/19/20
Posts: 612
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Re: SAB microparticles flying around, problem [Re: mad_dingo]
    #27006488 - 10/27/20 02:31 PM (3 years, 3 months ago)

Quote:

Maybe I'm not fully understanding what you say about the rack




The rack is a cross grid instead of horizontal metal bars. Naturally I cannot place any finger below the dishes to grab them on a cross grid. But the idea of sliding them over the edge of the rack like you do is good. could you show a picture of your kind of rack?

Quote:

How long had your SAB sat before you checked it with the torch?




First I spray the box and the table area with the towel, then I put the box on the table in its position for SAB work and I also spray inside it through the holes. The holes are kind of biggish (5 inches i think). I then let it sit 2-3 mins, proceed to do my work. I checked it after I was done working in it.

Quote:

Have you been have contam issues?



yea although work was being done with unsterilized dishes (didnt know until too late) so idk which part was inherent contams in the plates and which part sab contam.

thanks


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OfflineKmacmo
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Registered: 08/14/19
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Re: SAB microparticles flying around, problem [Re: coversall]
    #27006516 - 10/27/20 02:44 PM (3 years, 3 months ago)

Could be that air can never really be 'still' outside of a controlled environment, we just reduce the draft as much as possible to have minimal movement in the air. The SAB box will help trap the air that is inside it. (if you were to wave your hands around inside the box that would also disturb the air inside)
Spraying the air inside the SAB and in the room with some disinfectant spray will help kill anything in the air surrounding the SAB but it's not necessary.
Can't really prove this but just throwing my idea out there, I've never actually checked my SAB with a torch that's a great idea. But I don't usually get any contamination problems even sometimes I forget to close a window or two and it doesn't seem to affect anything for me.

3 minutes isn't really long enough to allow the air to settle down.


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OfflinePhony Phone

Registered: 09/19/20
Posts: 612
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Re: SAB microparticles flying around, problem [Re: Kmacmo]
    #27006530 - 10/27/20 02:50 PM (3 years, 3 months ago)

I see, thanks.
guess ill proceed and will update this thread regarding the dishes' success or otherwise failure. more setting down time added to the list of prerequisites.
:smirk:

edit: how long do you let yours sit and settle?

and 1 more thing, might seem stupid but whatever. After dish pouring you laeve them to cool down inside the sab, then you come back to wrap them up or do transfer work however you need to spray the sab again but the dishes are unwrapped and exposed now. What methods do you guys follow to work aroun this?


Edited by Phony Phone (10/27/20 02:59 PM)


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OfflineOne of Us
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Re: SAB microparticles flying around, problem [Re: Phony Phone]
    #27006550 - 10/27/20 03:02 PM (3 years, 3 months ago)

I leave my SAB in place, so I only let it settle a few minutes if at all. Just enough time for my scalpel handle to cool. I torch the whole length of it until its too hot to handle before putting it on its makeshift tin foil stand in the SAB. The room its in is pretty draft free too.

I think I remember seeing here, someone did the math and found the terminal velocity of spores, and figured it took about 30-40mins for a spore to travel from the roof to floor of sab... but I could be remembering wrong. 

I would say 20-30mins would be more than enough.


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OfflinePhony Phone

Registered: 09/19/20
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Re: SAB microparticles flying around, problem [Re: One of Us]
    #27006556 - 10/27/20 03:09 PM (3 years, 3 months ago)

Quote:

One of Us said:
I leave my SAB in place, so I only let it settle a few minutes if at all. Just enough time for my scalpel handle to cool. I torch the whole length of it until its too hot to handle before putting it on its makeshift tin foil stand in the SAB. The room its in is pretty draft free too.

I think I remember seeing here, someone did the math and found the terminal velocity of spores, and figured it took about 30-40mins for a spore to travel from the roof to floor of sab... but I could be remembering wrong. 

I would say 20-30mins would be more than enough.




Do you spray soapwater through the walls while the sab stays rooted in its usual position?
The spores from the roof surely do a lot of time to reach the bottom however the airborne bacteria and particles that fly around dont, because they are already everywhere including the agar level surface. so one has to either be super fast with the transfer and roll the dice or there must be a way to reaaally still up that air box. I haven't found a way around the second issue.


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OfflineOne of Us
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Re: SAB microparticles flying around, problem [Re: Phony Phone]
    #27006577 - 10/27/20 03:25 PM (3 years, 3 months ago)

I don't spray my SAB ever. I go in dry

I don't think bacteria "flies around" I think most bacterial contams in the SAB come from having unsterile tools or hands over open media. Maybe bumping the box can dislodge some from the roof of the sab.

But yeah, most of the waiting is for spores to settle, rather than bacteria.


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OfflinePhony Phone

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Re: SAB microparticles flying around, problem [Re: One of Us]
    #27006584 - 10/27/20 03:30 PM (3 years, 3 months ago)

Good to know. I was thinking more that bacteria is attached to the particles i spotted under light not literally flying around. :tongue2: But i guess your testimony of dry work working might mean that something else was going wrong wtih my batch.

How is your success rate innoculating petris and transfers etc?


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Invisiblecoversall
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Re: SAB microparticles flying around, problem [Re: Phony Phone]
    #27006606 - 10/27/20 03:42 PM (3 years, 3 months ago)

Mmmm I reckon the un-sterile dishes might be a bigger factor here.

tbh I've never thought of the SAB as a perfect piece of gear, it's some holes cut in a box at the end of the day.

My rack is pretty much identical to this thing



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OfflineOne of Us
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Re: SAB microparticles flying around, problem [Re: Phony Phone]
    #27006616 - 10/27/20 03:46 PM (3 years, 3 months ago)

Quote:

Phony Phone said:
Good to know. I was thinking more that bacteria is attached to the particles i spotted under light not literally flying around. :tongue2: But i guess your testimony of dry work working might mean that something else was going wrong wtih my batch.

How is your success rate innoculating petris and transfers etc?




I would say I have a 98%+ success rate, and 99% of my unsuccessful transfers are contaminated by mold rather than bacteria. I only really see bacteria on my germination plates


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OfflinePhony Phone

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Re: SAB microparticles flying around, problem [Re: One of Us]
    #27007541 - 10/28/20 02:11 AM (3 years, 3 months ago)

Ok figured it out.
Sprayed the box with soapwater while on its normal position. Then let it sit for several minutes with cling wrap covering the hole, 30 or 40 mins dont rememeber but i reckon 15 must be fine too.
So i went to recheck inside and the air was STILL. Nothing flying around.

Conclusion is, i probably created a draft at one point during my spraying sessions. The air got pushed in (probably when i sprayed the air around the box)  and the little dust got pushed in.
Lets hope my dishes will remain unscathed after this fieata.
Lesson learned.


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OfflineA.k.aM
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Re: SAB microparticles flying around, problem [Re: Phony Phone]
    #27007669 - 10/28/20 05:55 AM (3 years, 3 months ago)

The particles flying around after spraying the soap water is a good thing, they help pull spores out of the air as they fall to the ground.


Definitely keep your stuff raised up and try not to slide anything or bump the floor while you’re working cuz that’s where all the contams settle and it takes very little to whip them up.

I use a rack with a couple old plates stacked on top covered in iso soaked paper towel, then set the new ones on top of the stack.


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OfflinePhony Phone

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Re: SAB microparticles flying around, problem [Re: A.k.a]
    #27007734 - 10/28/20 07:12 AM (3 years, 3 months ago)

yea, ill try that, last session involved the plates sitting directly on the rack which was dripping wet. time will show if any damage was done this way.

also i further discovered that the "dust" particles i observed the other day were actually soap water droplets that had not settled and kept flying around with my hand movements.
Not that its safer for this to happen but it further decreases chances that my plates have been contam'd

and used saran wrap on the newer plates as they seemed too flimsy and fragile, didnt want to apply pressure using para

thanks to everyone who contributed to this thread :super:


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Offlineel gordo
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Re: SAB microparticles flying around, problem [Re: A.k.a]
    #27007754 - 10/28/20 07:25 AM (3 years, 3 months ago)

my advice is to build a small flowhood if u can, once u use a flowhood threres no turning back, a lot less hassle and mess each time u have to work plus 100% sure about cleanlines. Since i have one i feel like all the sab work ive done before was like trying to filter water with a sock full of sand when i could just put a chloride tab. may get similar results sometimes but the luck factor is always there if your procedure and skills are not very trained


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InvisibleTheMadHatter420
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Re: SAB microparticles flying around, problem [Re: el gordo]
    #27007779 - 10/28/20 07:42 AM (3 years, 3 months ago)

You could skip the SAB and buy or build a flow hood!


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