Primary cultures are difficult to transfect, especially neurons. We've made a generic reagent for primary cells (https://altogen.com/product/altofect-transfection-reagent/) and it's been validated for siRNA delivery. If you're using established primary cell cultures like Neuro2a, we've made kits for that as well (https://altogen.com/product/neuro-2a-transfection-reagent-neuroblastoma-cells-ccl131/). Toxicity in primary cells with generic reagents is pretty common, so I would look into pre-optimized reagents designed for primary cell cultures.

You may want to look at the attached paper.

Seems from different sources that Lipofectamine 2000 (invitrogen/Thermo) and jetPEI (Poly-plus) have been used successfully with synthetic siRNA.

if u want to silece your target gene  using RNAi  then u can also go for shRNA construct. shRNA transfection using lipofectamine 2000 as well as lipofectamine 3000 is easy. it has worked for me always. Also delivering a plasmid is easier than delivering an oligomer into cells. 

Hi Yuan,

I recommend NeuroMag which is based on Magnetofection technology it has been successfully used to transfect primary neurons (cortical, hippocampal, DRG, motor neurons, vagal afferent.....) You can see all citations here:

http://www.ozbiosciences.com/transfection-cell-specific/52-neuromag-neuron-transfection-magnetofection.html

In addition, here are few examples of direct link to publication for siRNA transfection in neurons

http://www.ncbi.nlm.nih.gov/pubmed/24027266

http://www.ncbi.nlm.nih.gov/pubmed/22731248

http://www.ncbi.nlm.nih.gov/pubmed/21940441

http://www.ncbi.nlm.nih.gov/pubmed/23585551

http://www.ncbi.nlm.nih.gov/pubmed/23620518

Good Luck

Olivier

thanks，guys！It is really helpful! I wonder Which is better regarding the transfection agent, 2000 or RNAiMAX? 

Lipofectamine 2000 works fine, but neuron density is key and you might need to increase it compared to normal cultures. Efficiency with siRNA tends to be better than with plasmids, but nothing compared to a cell line...

For siRNAs I have usually had better luck with RNAiMAX than 2000.

Thanks for your advice,Jonathan. What is your working concentration of RNAiMAX in your transfection system?  How do you minimize the toxictity to your primary culture?

You can please follow the links below to obtain valuable informations:-

1. http://www.ncbi.nlm.nih.gov/pubmed/20709387

2. http://www.ncbi.nlm.nih.gov/pubmed/16472690

3. http://www.nature.com/mtna/journal/v2/n12/full/mtna201365a.html

4. http://www.jneurosci.org/content/24/45/10040.full

Thanks,Sandip Chakraborty! It is really helpful~

Hi, Yuan. Did you find other good ways to deliver the siRNA to primary cultured neuron? I also tried RNAiMax, my neuron all died too. So wondering, how did you optimize your system or turn to other method? Many thanks!~~~

Mengying, I tried other methods to deliver siRNA, but they all failed in my system, Now I am using shRNA to knockdown my target gene. 

Yuan, thanks a lot. Now I am also thinking to use shRNA. Good luck to both of us.

Go to Advirna

Primary cultures are difficult to transfect, especially neurons. We've made a generic reagent for primary cells (https://altogen.com/product/altofect-transfection-reagent/) and it's been validated for siRNA delivery. If you're using established primary cell cultures like Neuro2a, we've made kits for that as well (https://altogen.com/product/neuro-2a-transfection-reagent-neuroblastoma-cells-ccl131/). Toxicity in primary cells with generic reagents is pretty common, so I would look into pre-optimized reagents designed for primary cell cultures.