Hi Abdelraheim Attaai ,

Interesting! I was also involved in a study on scopolamine-induced AD model with focus on changes in hippocampal (DG) neurogenesis.

To answer your questions:

1. In my opinion, you need more samples to conduct both tests, especially for western blot which needs more specimen from hippocampus. I recommend using hippocampi of both hemispheres in different rats from your various groups.

Try to dissect and have a clean extraction of hippocampi though they can be far more easily extracted from the brain compared to other brain regions.

2. It depends on the period of time that you would like to store your specimen, and also the type of molecular experiments in which you would need to perform after WB or ELISA to interpret your results. But I think that PBS would be sufficient for your tests.

Article The Unfolded Protein Response Is Activated in Pretangle Neur...

https://science.sciencemag.org/content/290/5493/985.short

Article Synchronization of GABAergic interneuronal network in CA3 su...

HEPES can be used as a wash buffer.

3. If you need to keep your samples for not too long, then homogenization would be OK in advance (in a controlled temperature).

kindly, go to this paper

https://www.sciencedirect.com/science/article/abs/pii/S0024320519301407

Hi Abdelraheim Attaai,

I think it's totally possible. I use mouse HC from only one hemisphere to run WB using c400 Imaging System (Azure Biosystems). So, as rat HC is much bigger, it should be enough to do both.

I use dry ice-cold saline for washing and keep the selected brain structures in -80°C after dissection until further processing (without any buffer). Then, cut them on dry ice and homogenize using ice-cold 1 × Cell Lysis Buffer.

Good luck!