Monday, June 20, 2011

Transfection/Infection of Primary Neurons

Gene Expression Analysis of Neurons is an important tools in basic research and the study of neuropathologies. At the Neuromics' blog: "siRNA, DsiRNA and Plasmid Transfection Efficiency", I have posted many examples of successful tarnsfection of primary neurons and related cells using both our Transfection Kits/Reagents and others.

The other puzzle piece for these studies is having a fresh, pure and easy to use source of cells. Here, Neuromics has many options. These primary neurons and neural progenitors are widely referenced in key publications. Applications referenced include: transfection, pharmacology, electrophysiology, immunocytochemistry, and neuronal development studies.

This posting features infection of our e18 Primary Rat Combined Hippocampus, Cortex, and Ventricular Neurons using Nipah virus related components and HeV pseudotyped virions

Aparna Talekar, Antonello Pessi, and Matteo Porotto. Infection of primary neurons mediated by Nipah virus envelope proteins: Role of host target cells in antiviral action. J. Virol. doi:10.1128/JVI.00452-11.

Abstract: We have previously described heterotypic peptides from parainfluenza virus that potently inhibit Nipah virus in vitro, but are not efficacious in vivo. By contrast, our second-generation inhibitors, featuring a cholesterol moiety, are also efficacious in vivo. The difference between in vitro and in vivo results led us to investigate the basis for this discrepancy. Here we compare the activity of the compounds in standard laboratory cells and in cells relevant to the natural tropism of Nipah virus, i.e. primary neurons, and show that while our first generation inhibitors are poorly active in primary neurons, the cholesterol-conjugated compounds are highly potent. These results highlight the advantage of evaluating antiviral potency in cells relevant to natural host target tissue.

Customer Data: Transfection of functional HeV glycoproteins and infection with HeV pseudotyped virions.In order to establish the feasibility of carrying out the proposed experiments in primary neurons, we show (figure ) that our assays are amenable to use in primary neurons. In the experiment, Combined Hippocampus, Cortex, and Ventricular -E18 (Neuromics) were plated, and at 3 days were transfected with plasmids encoding HeV G/F as well as YFP. On the following day, these cells were infected with HeV or VSV pseudotyped viruses bearing RFP. In the figure, (A) the phase contrast photos show the differentiated neurons; (B) upon excitation for RFP, the red fluorescence indicates neurons infected by HeV pseudotyped virions; (C) upon excitation for YFP, and the green fluorescence shows the efficiency of transfection in neurons. This experiment indicates that the proposed experiments can be carried out in primary neurons, which are transfectable and infectable in our systems, and thus supports all the proposed aims. Data Courtesy of Dr. Matteo Porotto, Weill Cornell Medical College. Larger Image

We will continue to keep you updated.

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