Optimal density to transfect adult fibroblasts using Epi5 Vector??

02 March 2022 1 5K Report

We are using the Neon Transfection system with Epi5 Vectors. The fibroblasts will be at passage 3 when transfected and they are from an adult donor. After transfection they will be placed in N2B27+100bFGF for 15 days and then switch to Tesr-E8. What would be the optimal seeding density for each well of a 6 well plate? Additionally is 1650V appropriate to transfect adult fibroblasts? The protocol on the website is optimized for fetal fibroblasts.

Shin Murakami

https://www.atcc.org/-/media/product-assets/documents/protocols/cell-biology/dermal-fibroblasts-on-letterhead-final.pdf?rev=b0345bd517da4e9880bf52dbe819b567

https://tools.thermofisher.com/content/sfs/brochures/FAQs_Epi5_Episomal_iPSC_Reprogramming_Kit.pdf

https://altogen.com/Fibroblast_Transfection_Protocol.pdf

Article Optimization of episomal reprogramming for generation of hum...

Various protocols are available above. ATCC says "Cell density should be 50-80% confluent" but you could explore a series of confluencies depending on your experimental condition. Good luck.

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