Author Topic: BDNF IHC?  (Read 7210 times)

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Offline hbrenhouse

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BDNF IHC?
« on: November 13, 2003, 04:39:48 PM »
Hi there,

I would like to start an IHC stain for BDNF, but I've heard it is extra-tricky.  Does anyone have any experience with this in rat brain?

Thanks for any advice
Heather

BDNF IHC?
« on: November 13, 2003, 04:39:48 PM »

Offline Jossy

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BDNF IHC?
« Reply #1 on: June 23, 2004, 06:33:53 PM »
Hi, I've realized a IHC for NGF, and works well, maybe my protocol works for you too. :D
(criostate slides of 40 micrometers)

Wash 15 min in PBS (0.1M, pH 7.4) at RT
Incubate 15 min in 0.3% H2O2 (in PBS) at RT
Wash 5 min in PBS at RT
Incubate 3 times in EtOH 50% (in PBS), 10 min each, at RT
Wash 15 min in PBS, at RT
Incubate 1 hr in PBS + 0.2% Triton x-100 / 5% Normal Serum at RT
Incubate with primary antibody (1:500), in PBS + 0.2% Triton x-100 / 2% Normal Serum at 4 C O.N.
Wash 3 times in PBS , 15 min each, at RT
Incubate with secondary biotinylated antibody (1:500), in PBS + 0.2% Triton x-100 / 0.1% NS, 2 hrs at RT
Wash 3 times in PBS , 15 min each, at RT
Incubate one and a half hour with ABC Kit (1:110) diluted in PBS, at RT (mix 30 min before use)
Wash 3 times in PBS , 15 min each, at RT
Incubate in 0.04% DAB + 0.01% H2O2 + NiCl2 0.06%, 10 min at RT (discard to chlorine solution)
Wash 3 times in PBS , 15 min each, at RT (discard to chlorine solution)

Offline hbrenhouse

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Thanks, Jossy!
« Reply #2 on: June 24, 2004, 09:15:50 AM »
Thanks, Jossy!  I've heard that there are few, if any, good BDNF antibodies out there, but if I find one I will give your protocol a try.  Thanks again

Offline SGlerup

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Re: BDNF IHC?
« Reply #3 on: August 10, 2012, 04:56:17 PM »
We have used an antibody against the proform of BDNF (proBDNF) that works very well for IHC on mouse embryo and brain sections.
It is a rabbit polyclonal from Alomone.

I have reviewed it on www.pabmabs.com

A postdoc from our lab has also used BDNF mAb #9 from Yves Barde's lab (Matsumoto et al. 2008 Nat Neurosci) with success on primary hippocampal neurons treated with KCl.

Offline AlexG

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Re: BDNF IHC?
« Reply #4 on: January 14, 2014, 02:52:16 PM »
I don't see any information about how the animals were sacrificed/perfused and how the tissue was post-fixed. I was wondering if you could supply this information? Thank you so much. Looking forward to hearing back.

- Alex

Re: BDNF IHC?
« Reply #4 on: January 14, 2014, 02:52:16 PM »