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autoclaving


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#1 lyok

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Posted 30 August 2009 - 11:38 PM

Dear all,

I have the following, maybe strange question: do you lose a lot of fluid when autoclaving a bottle of fluid?


I am going to make a dilution and since I can not autoclave 1 main component of my mixture I was going to autoclave all the rest and then simply add the substance I can not autoclave, but my concern now is that when I make this solution to be autoclaved, I take exactly how much I need. So that I can simply add the amount of non autoclaved substance to it. But will this calculation still be ok? Or will I lose a lot of the fluid and thus need to repipette the autoclaved substance to check if it is still ok ?

ex: I need to ad 50ml of substance B (can not be autoclaved) and I put in in 250ml of the solution I do autoclave, but when I autoclave this 250ml, will I lost a lot of it during the autoclaving?

#2 lsek

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Posted 30 August 2009 - 11:47 PM

You can try covering the bottle cap with aluminium foil (that works for me by limiting "fluid loss" during autoclaving). Normally, I wouldn't really be too worried about minute differences in the solution preparation.

Bests

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#3 Stephan

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Posted 31 August 2009 - 12:02 AM

Ratios are always important. If you measure out exactly the right amount of substance (250ml) and then autoclave, you will always loose fluid through evaporation. If you don't want to do this then you can seal the bottle, but this will possibly explode the bottle due to high pressures (unless you have something special for this).

I would really suggest simply making more than enough and re-measuring after autoclaving under a lamina flow. This is not really much more work, is it? Rather be sure of your volumes.

What are you making by the way?

#4 lyok

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Posted 31 August 2009 - 12:20 AM

Ratios are always important. If you measure out exactly the right amount of substance (250ml) and then autoclave, you will always loose fluid through evaporation. If you don't want to do this then you can seal the bottle, but this will possibly explode the bottle due to high pressures (unless you have something special for this).

I would really suggest simply making more than enough and re-measuring after autoclaving under a lamina flow. This is not really much more work, is it? Rather be sure of your volumes.

What are you making by the way?



Nothing really special, just a CaCl2 solution; but since I can not autoclave the CaCl2, I'll add this later on.

Its just to prepare competent cells.

I need a 0.1M solution of CaCl2, but I think if I lose some of my water and glycerol it doenst really matter a lot; then I"ll have 0.12 or something like that in stead of 0.1.

I think its not that important now.

But I was thinkin about it in the general sence , for the future , when I need more precise solutions.

#5 leelee

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Posted 31 August 2009 - 01:33 AM

Are you losing heaps or just a little? I have noticed heaps of people in my department loosen the lids of bottles way too much when autoclaving, and then their solution boils over and they lose several mls. You really only need to loosen about 1/4 turn, just to release the pressure that builds up- and then you shouldn't lose too much.

#6 bob1

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Posted 31 August 2009 - 01:57 PM

You could try filter sterilisation. A 0.2 micron (um) filter will work for anything apart from viruses and mycoplasma

#7 lab rat

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Posted 31 August 2009 - 02:08 PM

Hi Lyok,

If you use a bottle larger than the amount of liquid, and set it in a pan with a little water during the liquid cycle, you will minimize your chances of boilover.

Your loss by evaporation will be minimal--the steam from the autoclave will prevent too much from escaping the bottle. Just loosely set the cap (too tight, and you won't get it back off) after autoclaving and let cool.

Regards,

lab rat
42..."An immutable fixed-precision number of unlimited magnitude." <a href="http://en.wikipedia....amming_language)" target="_blank">http://en.wikipedia....amming_language)</a>, accessed 25June2009.

#8 GeorgeWolff

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Posted 31 August 2009 - 03:33 PM

Agree - and lee lee is incorrect, you don't get "boil over" by loosening the cap too much.

#9 swanny

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Posted 31 August 2009 - 05:41 PM

As an alternate to the hood, you could always try weighing the bottle before and after autoclaving and add the difference.
Heart disease kills more women than breast cancer, but heart attack symptoms differ from men's symptoms. Get to know your heart... it could save your life.

#10 leelee

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Posted 31 August 2009 - 05:57 PM

Maybe I am....... don't know. All I know it that when some members of the department who were experiencing problems loosened their caps less, the loss of liquid they were experiencing stopped- maybe less evaporation??

#11 eberthella

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Posted 31 August 2009 - 07:05 PM

Think about it - why should a (loose) cap loosened more lead to more boil over?

lab rat - I understand the "bottle larger than the amount of liquid" and I'm sure autoclaving a bottle smaller than the amount of liquid would be challenging. I just can't see many folks trying it. Have you?

#12 leelee

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Posted 31 August 2009 - 09:31 PM

What exactly is you reason for being on this forum, eberthella?? From what I can see, all you seem to do is go around insulting people and trying to cause trouble. The whole reason for the forum is for us to help each other out. We don't always have the right answers, and we don't always express what we are trying to say in the clearest and most accurate way. But at least we are trying to be helpful. Which is a heck of a lot more than can be said for you.

#13 eberthella

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Posted 01 September 2009 - 05:07 PM

Sorry you're so sensitive. I could have just said you were wrong like George but merely asked you to think. Sorry if asking you to think is so offensive . I'll not make such a bizarre request again.

#14 leelee

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Posted 01 September 2009 - 05:49 PM

I actually wasn't talking about what you said to me, I was talking about your comment to labrat. It was abundantly clear to me (and I'm sure most others) what labrat was trying to say, and yet you still felt the need to be a smart-ass about it.

#15 eberthella

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Posted 01 September 2009 - 06:10 PM

Not sure anything is abundantly clear to you, leelee, so let me help you. A vessel smaller than it's contents is an anolomy and lab rat is an adult. He or she doesn't need a person as inexperienced in the lab and selfconscious as you to speak for them. Understand?




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