Re: [Histonet] low signal for long post fixation

2017-03-23 Thread Ray via Histonet 
Mariela, 

(1) If you are looking at enzyme activity of a LacZ transgene to localize with 
X-gal staining, I found the situation really a problem with extended fixation 
times of vibratome sections (like you with controls or test article). Have 
never been able to "retrieve" that kind of staining. Maybe someone out there 
knows the trick. Especially bad in low-level integration of transgene. 
(2) I have though gone after the enzyme with just traditional IHC to 
B-galactosidase (there are good antibodies available. And as Rene said, 
increase your antigen retrieval which will (might, should) bring back the 
antigenic sites for IHC. But for the enzyme activity, I've never had much luck 
at that. 

Ray Koelling 
Faculty lecturer, WWAMI, UW School of Medicine, Spokane, WA 

- Original Message -

From: "Mariela Chertoff via Histonet" <histonet@lists.utsouthwestern.edu> 
To: histonet@lists.utsouthwestern.edu 
Sent: Thursday, March 23, 2017 6:28:24 AM 
Subject: [Histonet] low signal for long post fixation 

Dear all, 

I have mouse brain tissue perfused with PFA 4% and post fixed in formol 10% 
for periods between 2 month and 1 year. I cut them on vibratome (30um) and 
I made inmunohistochemistry and beta galactosidase to check senescence. 
My problem is that I have less signal on positive controls if postfixation 
on formol was longer. 

There is a way to solve this problem? 

Thanks in advance for your reply 

Mariela Chertoff, PhD 
Laboratorio de Neuroepigenetica - QB75 
Departamento de Química Biológica 
Facultad de Ciencias Exactas y Naturales - UBA 
Ciudad Universitaria Pabellón II Piso 4 
Ciudad Autónoma de Buenos Aires 
C1428EGA - Argentina 
Tel: 54 11 4576-3300/09 - Int. 221 
email:marielachert...@gmail.com 
marielachert...@qb.fcen.uba.ar 
<mariela.chert...@uab.cat> 
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Re: [Histonet] low signal for long post fixation

2017-03-23 Thread Rene J Buesa via Histonet 
Perhaps your only solution is to increase HIARRené 

On Thursday, March 23, 2017 9:39 AM, Mariela Chertoff via Histonet 
 wrote:
 

 Dear all,

I have mouse brain tissue perfused with PFA 4% and post fixed in formol 10%
for periods between 2 month and 1 year. I cut them on vibratome (30um) and
I made inmunohistochemistry and beta galactosidase to check senescence.
My problem is that I have less signal on positive controls if postfixation
on formol was longer.

There is a way to solve this problem?

Thanks in advance for your reply

Mariela Chertoff, PhD
Laboratorio de Neuroepigenetica - QB75
 Departamento de Química Biológica
Facultad de Ciencias Exactas y Naturales - UBA
Ciudad Universitaria Pabellón II Piso 4
Ciudad Autónoma de Buenos Aires
C1428EGA - Argentina
Tel: 54 11 4576-3300/09 - Int. 221
email:marielachert...@gmail.com
marielachert...@qb.fcen.uba.ar

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[Histonet] low signal for long post fixation

2017-03-23 Thread Mariela Chertoff via Histonet 
Dear all,

I have mouse brain tissue perfused with PFA 4% and post fixed in formol 10%
for periods between 2 month and 1 year. I cut them on vibratome (30um) and
I made inmunohistochemistry and beta galactosidase to check senescence.
My problem is that I have less signal on positive controls if postfixation
on formol was longer.

There is a way to solve this problem?

Thanks in advance for your reply

Mariela Chertoff, PhD
Laboratorio de Neuroepigenetica - QB75
 Departamento de Química Biológica
Facultad de Ciencias Exactas y Naturales - UBA
Ciudad Universitaria Pabellón II Piso 4
Ciudad Autónoma de Buenos Aires
C1428EGA - Argentina
Tel: 54 11 4576-3300/09 - Int. 221
email:marielachert...@gmail.com
marielachert...@qb.fcen.uba.ar

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