Dear Devon,

We do have two options for transfecting SH-SY5Y that work very well for 'classic transfections": either lipofection or Magnetofection*** (you can refer to our citation database in the link below).

Concerning CRISPR/Cas9, it is important to take into consideration what you are about to transfect to give you the best answer:

• Cas9 endonuclease can be delivered via a plasmid, mRNA, protein, or virus.
• sgRNAs can be coded by a plasmid, a viral vector, or directly transported.
• moreover, some plasmids or virus can encde for the two molecules.

So, depending on the material you have for CRISPR/Cas9 knock out, you will have to use different tool (refer to the link below) . As specialists of transfection/transduction, we do have tools for any biomolecule to deliver and I would be very glad to asist you in setting up your protocol.

You can contact me directly via ResearchGate or at: [email protected]

good luck with your experiment,

best regards,

Cedric

*** The Magnetofection technology uses a magnetic field to attract and concentrate complexes of magnetic nanoparticles and nucleic acid onto the cell surface as demonstrated by Grześkowiak BF et al, Pharm Res. 2014 Jul 18. (http://www.ncbi.nlm.nih.gov/pubmed/25033763).

http://www.ozbiosciences.com/module/citationfinder/default

http://www.ozbiosciences.com/content/58-transfection-reagents-for-genome-editing

We've had some good success transfecting these cells with Lipofectamine 3000. You could also add a DNA ligase inhibitor to the media as you transfect and that may help. We've had about 15-20% transfection efficiency using both. 

We use Lipofectamine LTX and Plus Reagent to transfect SH-SY5Y cells.

Thank you everyone!

Are all these suggestions in the 20% transfection efficiency? what about the ability of transfection two plasmids, like Devon will need to do i.e.: CRISPR/Cas9

won't this lower the efficiency even more?

Dear Jessica,

you are definitively right, the efficiency of the transfection is inversely related to the number of plasmid. I have recently asnwered to a post (in the link below) which you can refer to.

This is why we do recommend magnetofection as this method is really efficient and present a high compaction level of nucleic acid allowing multiple or large plasmid transfections.

would you like to try it; please do not hesitate to contact me;

best regards,

Cedric

https://www.researchgate.net/post/Has_anyone_tried_to_transfect_four_plasmids_4000_bp_7000bp_all_at_the_same_time_to_HEK_and_HMECs

Anyone transfected SH-SY5Y with dual plasmid systems using Attractene. I harvested cells after 24 hours of transfection. I did not observe any messy sequence in my Sanger sequencing result.

Neon electroporation with RNP could be an option to try for SH-SY5Y cells. They have a better efficiency than the classical plasmid transfection.

Devon, did you have any luck using CRISPR-Cas9? I've been only seeing HEK cells being used as an alternative to SH-SY5Y cells. I was looking forward to knockout a gene in this cell type, but I've been wondering the same thing as you. Did you try and have any feedback? It could be very useful to me. Thank you.

Using CRISPR/Cas9 in SH-SY5Y cells has been explored in various research projects with the aim of understanding genetic functions and modeling neurological diseases. SH-SY5Y cells are a human-derived cell line often used in neuroscience research because of their ability to differentiate into neuron-like cells, making them a valuable model for studying neurodegenerative diseases, neuronal function, and drug screening.

Researchers have successfully applied CRISPR/Cas9 technology to edit genes in SH-SY5Y cells for various purposes, such as creating models of neurological diseases, studying gene function, and investigating therapeutic targets. For example, CRISPR/Cas9 has been used to introduce mutations relevant to Parkinson's disease, Alzheimer's disease, and other neurodegenerative conditions into these cells. This enables the study of the pathological mechanisms of these diseases and the testing of potential therapeutic interventions.

The success of using CRISPR/Cas9 in SH-SY5Y cells depends on several factors, including the design of the guide RNAs (gRNAs), the efficiency of the CRISPR/Cas9 delivery method (e.g., plasmid, viral vectors), and the selection and validation of edited cells. Researchers typically report modifications in the efficiency of gene editing, challenges in achieving high specificity without off-target effects, and strategies for optimizing the differentiation of edited SH-SY5Y cells into neuron-like cells to study functional outcomes.

If you're planning a CRISPR/Cas9 project with SH-SY5Y cells, reviewing the latest literature in this area is crucial for understanding the specific strategies, efficiencies, and challenges reported by other researchers. This can help in designing your experiments and anticipating potential obstacles. Are you looking for information on specific experimental designs or outcomes related to CRISPR/Cas9 editing in SH-SY5Y cells?

l This protocol list might provide further insights to address this issue.