how to degas small volume liquid ( microliter )
Frank
Hello, anyone knows how to degas a very small volume?
My problem is like this: I have to work with very small sample
quatilities, usually microliter range. Basicly, my samples are mostly
water/buffer with genomic/proteins molecules dissolved in it. I inject
the sample to a 5 microliter chamber and heat up to 45 degree for the
reaction. But I found that bubbles start to form at the sidewalls of
the chamber. I think this may due to the solibility decrease of air in
water at the high temperature. I cannot use normal vacuum degas method
because it will evaporate water and change the concentration of my
sample. I'm hesitate to use ultrasonic because I'm afraid it may do
sth bad to DNAs in the sample... I'm thinking of heating the sample to
45C before the injection. But for such a small volume, after pipetting
and injecting, I'm afraid that it will fall back to the room
temperature.
Any other smart option? Thank you very much for your help!
Frank
Jim Kelley
increasing the pressure.
Mark Thorson
Frank wrote:
> Any other smart option? Thank you very much for your help!
Bubble helium gas though it?
dlzc@aol.com (formerly)
Dear Frank:
"Frank" <youn...@yahoo.com> wrote in message
news:b6avj...@enews1.newsguy.com...
I know nothing about your procedure. Can you increase the pressure on the
sample by a couple of psi (aka. 0.2 bar) or so? This will help the
counteract the change in solubility...
David A. Smith
Chris Lee
You degas HPLC mobile phases by bubbling helium through them for a short
time. Helium is more-or-less insoluble in most liquids.
Bearing in mind the small volume, perhaps you could just replace the air
with helium, with a little vortexing if necessary.
Regards
"Frank" <youn...@yahoo.com> a crit dans le message de news:
b6avj...@enews1.newsguy.com...
..
Uncle Al
Work with it under helium. Helium has very low solubility in water.
Humidify your helium by bubbling through degassed or purged water.
Pressurize before heating - though it may foam on cooling anf breaking
the pressure.
--
Uncle Al
http://www.mazepath.com/uncleal/
(Toxic URL! Unsafe for children and most mammals)
"Quis custodiet ipsos custodes?" The Net!
Mark Thorson
Frank wrote:
> Any other smart option? Thank you very much for your help!
Bubble helium gas through it (called "sparging").
Helium has low solubility in aqueous solutions,
but the gas in the solution will happily
diffuse into the helium.
TomB
You can degas a liquid by the law of physics that the solubility of a
gas in a liquid decreases as the temperature increases and becomes
zero at the boiling point of the liquid. If you use a vacuum with very
specific temperature control, you can stop as soon as the fraction is
no longer gas and changes over to the liquid. This will not change the
concentration of the sample. The only other suggestion would be if
possible, to degas before the protiens are dissolved in the liquid.
Frank
for example, from internet I got:
15ml air dissolve in 1L water at 1atm at rt.(say 20c).
now the question is
15ml air dissolve in 1L water at XX atm at 45 C.
how to do the calculation? which formula should I use?
Thanks a lot!
Frank
Jim Kelley <jwke...@uci.edu> wrote in message news:<3E89D051...@uci.edu>...
TomB
youn...@yahoo.com (Frank) wrote in message
news:<b6avj...@enews1.newsguy.com>...
What problems are the bubbles causing? I think to bubble helium
through the sample will remove as much of the liquid as careful vacuum
degas. I also don't think you will have any problem if you heat to 45
degree before you inject if you will keep agitation to a minimum.
JuSu
> But for such a small volume, after pipetting
> and injecting, I'm afraid that it will fall back to the room
> temperature.
So let it fall back to room temperature. Re-absorption of gases is not
instantaneous.
How absolute do you really need the degassing to be?
If you degas water by boiling and let it cool down, I believe the
amount of gasses that have time to become absorbed in the liquid is
smallish.
If your needs for purity are high, other solutions given are more
applicable.
//JuSu
Frank
Thanks a lot for all the suggestions!!!
I appologize for posting this question around. I was really frustrated
with the bubbles in the tiny tiny tiny volume! I have to deal with 10
microliter. I got the sample from other labs and that's all I have. It
only occupies tiny bit bottom of an eppy tube. From all the
suggestions, it looks like degasing with humidified helium would be
the easiest way to give it a try if I can get helium easily.
Here is my proposed setup:
helium goes into a bottle with degassed water through a tube, make
sure helium bubble through this water. Then a tube connects the
outcoming helium from the bottle to the eppy tube and use a needle to
introduce helium bubble to the 10 microlitter sample. Since helium is
pre-humidified, the bubbles are saturated with water vapor so they
won't carry H2O away from my 10 microliter sample no matter how long I
carry out this process, am I correct?
Hope this works. Another option is to try pressurizing the chamber.
I'm not sure how well my chamber can hold the presure and how much
presure I should exert. How to calculate the presure needed to prevent
air from coming out of a 45C solution? Which formula should I use?
Thanks a million to all of you!!!
Frank
Lucius Chiaraviglio
youn...@yahoo.com (Frank) wrote:
>Here is my proposed setup:
>helium goes into a bottle with degassed water through a tube, make
>sure helium bubble through this water. Then a tube connects the
>outcoming helium from the bottle to the eppy tube and use a needle to
>introduce helium bubble to the 10 microlitter sample. Since helium is
>pre-humidified, the bubbles are saturated with water vapor so they
>won't carry H2O away from my 10 microliter sample no matter how long I
>carry out this process, am I correct?
Theoretically yes, but in practice, you can't get away with this
forever. Unless you have insanely good temperature control, your helium
humidification bottle will be at not exactly the same temperature as your
sample, and thus the humidified helium will either be not quite saturated
or slightly supersaturated. So don't go crazy with your sparging -- just
experiment a little with fake samples until you determine how much you
need to do to prevent bubble problems.
--
Lucius Chiaraviglio
Approximate E-mail address: luci...@chapter.net
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