Cloudy TBS solution.

[Ved Sharma]

Hi,

I'm having trouble keeping 1X TBS solution clear/transparent. It
becomes cloudy after a couple of days. I make it by diluting 10X
TBS solution and then adjusting the pH to 7.3. I don't autoclave
after that. Is that the reason why it gets cloudy?

Thanks!

Ved

----------------------------------
Ved P. Sharma
Graduate Student
Department of Chemical Engg
University of Florida
Gainesville, FL-32611
-----------------------------------

[GysdeJongh]

"Ved Sharma" <ve...@ufl.edu> wrote in message
news:mailman.554.114471...@net.bio.net...

> Hi,
>
> I'm having trouble keeping 1X TBS solution clear/transparent. It becomes
> cloudy after a couple of days. I make it by diluting 10X TBS solution and
> then adjusting the pH to 7.3. I don't autoclave after that. Is that the
> reason why it gets cloudy?

Mold's I guess
Make a fresh 1x dilution each day from the 10x TBS solution , or keep the 1x
TBS in the refrigerator for a few days if that is more convenient.Keep the
1x TBS always on ice while working with it.Think of 1x TBS as milk , the
bugs like it :(

The mold's and other bugs don't grow in the 10x Solution because the osmotic
pressure is way to high for them.You can keep the 10x TBS on your bench at
rt. I always filter sterilise the 10x TBS and treat it with care.

hth
Gys

[Jayakumar, R]

You don't need to autoclave TBS if you are going to use it only for
westerns, though filtering it is a good habit. How do you adjust the
pH?? I mean what acid do you use?

Jai

-----Original Message-----
From: methods...@oat.bio.indiana.edu
[mailto:methods...@oat.bio.indiana.edu] On Behalf Of Ved Sharma
Sent: Monday, April 10, 2006 3:56 PM
To: met...@magpie.bio.indiana.edu
Subject: Cloudy TBS solution.

Hi,

I'm having trouble keeping 1X TBS solution clear/transparent. It
becomes cloudy after a couple of days. I make it by diluting 10X
TBS solution and then adjusting the pH to 7.3. I don't autoclave
after that. Is that the reason why it gets cloudy?

Thanks!

Ved

----------------------------------
Ved P. Sharma
Graduate Student
Department of Chemical Engg
University of Florida
Gainesville, FL-32611
-----------------------------------

_______________________________________________
Methods mailing list
Met...@net.bio.net
http://www.bio.net/biomail/listinfo/methods

This email message may contain legally privileged and/or confidential information. If you are not the intended recipient(s), or the employee or agent responsible for the delivery of this message to the intended recipient(s), you are hereby notified that any disclosure, copying, distribution, or use of this email message is prohibited. If you have received this message in error, please notify the sender immediately by e-mail and delete this email message from your computer. Thank you.

[Ved Sharma]

Thanks for your reply.

I'm using TBS for immunohistochemistry and I adjust the pH by HCl.
I never tried filtering it because when I prepare the solution, it
looks very clear. Can it be a bacterial contamination which
develops after a couple of days? Although it's hard for me to
imagine if bacteria can live on a "Tris base-NaCl" diet.

Ved

On Mon Apr 10 21:55:00 EDT 2006, "Jayakumar, R"

[Nick Theodorakis]

Ved Sharma wrote:
> Thanks for your reply.
>
> I'm using TBS for immunohistochemistry and I adjust the pH by HCl.
> I never tried filtering it because when I prepare the solution, it
> looks very clear. Can it be a bacterial contamination which
> develops after a couple of days? Although it's hard for me to
> imagine if bacteria can live on a "Tris base-NaCl" diet.
>

You'd be surprised at what they grow in.

Does it smell funny when you see the turbidity? It's not always a sure
thing, but if it smells you definitely have contamination.

Nick

--
Nick Theodorakis
nick_the...@hotmail.com
contact form:
http://theodorakis.net/contact.html

[Christian Praetorius]

Ved Sharma <ve...@ufl.edu> wrote:

>develops after a couple of days? Although it's hard for me to
>imagine if bacteria can live on a "Tris base-NaCl" diet.

Its always astonishing in what conditions bacteria and fungi can grow
and survive. I would filter or autoclave the buffer.

Christian

--
[X] <-- nail here for new monitor

[Dr Engelbert Buxbaum]

Ved Sharma wrote:

> I'm using TBS for immunohistochemistry and I adjust the pH by HCl.
> I never tried filtering it because when I prepare the solution, it
> looks very clear. Can it be a bacterial contamination which
> develops after a couple of days? Although it's hard for me to
> imagine if bacteria can live on a "Tris base-NaCl" diet.

Probably either bacteria or fungi. Why not simply look at them in the
microscope?

By the way, keeping solutions in the fridge is good practice, as is the
addition of antibacterials like sodium azide (don't use if you detect
with peroxidase) or timerosal.