Hello, everyone!
I'm going to start a project with single-cell ATAC-seq for analysis of open chromatin regions from freeze tissue samples.
I'm lost about the nuclei concentration.
If I made a nuclei isolation protocol from the human brain freeze tissue sample (hippocampus), what would be the nuclei mean number from these?
2000 nuclei? 5000 nuclei?
There is any paper about that?
Thank you!
Hello Jaqueline,
I would follow the work of Sebastian Preissl (now at UCSD). He has much experience on isolated nuclei sequencing and should provide the guidance you need in his publications. He recommends the far-red DNA dye DRAQ7 to label the nuclei to sort them free of debris, etc. He gave an excellent webinar recently (see Selectscience for a recording of this.
Regards and good luck,
Roy
Check out the single nucleus isolation kit from Invent Biotechnologies. It is designed for specifically for neuronal tissues and works well with frozen tissues. The filter they use does a good job of separating myelin from the isolated nuclei to eliminate contamination and give a high yield. https://inventbiotech.com/products/minute%E2%84%A2-single-nucleus-isolation-kit-for-neuronal-tissues-cells-20-preps?_pos=1&_sid=1ba7889f6&_ss=r
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