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Transfection Reagent for HCN-1A Cells (Brain Neuronal Cells, CRL-10442)
Two component formulation enhances lipid mediated transfection efficiency
Optimized easy-to-use transfection protocol provided for transfection of siRNA, DNA, mRNA, and microRNA
Kit includes Transfection Enhancer reagent and recommended transfection protocol
High transfection efficacy in the presence of serum
Expand your RNAi application with a reagent optimized for delivery of both siRNA and plasmid
Reproducible transfection results
Download PowerPoint presentation for HCN-1A cells transfection kit: [PPT]
Developed and manufactured by Altogen Biosystems
Reagent exhibits at least 76% transfection efficiency of siRNA delivery. Transfection efficiency was determined by qRT-PCR.
Transfection Protocol and MSDS:
Download Altogen Biosystems HCN1A Transfection Protocol: [PDF]
Download MSDS: [PDF]
HCN-1A Cell Line:
A number of studies have demonstrated the successful application of RNAi in primary cell cultures and non-neuronal cell lines; however the use of gene silencing in neuronal cell types has been notoriously difficult because of several technical and biological limitations. Neurons have been considered the most resistant to RNAi. Methods to consider when performing nucleic acid delivery experiments with neuronal cultures are: electroporation, microinjection, viral-based methods, and certain chemical transfection reagents. Although physical delivery methods appear to be efficient for siRNA-induced studies where transient modification of gene expression is sufficient, these methods do not appear to be adequate for behavioral studies where long term alteration of gene expression may be necessary. The viral-based siRNA expression systems appear to be most applicable for both Central Nervous System (CNS) gene therapy and basic neurobiological studies. Lentiviral vectors have been reportedly used successfully for the delivery of short hairpin RNA (shRNA), a precursor of siRNA, into primary neurons to induce RNAi.
The HCN-1A cell line is a human cortical neuronal line derived with tissue taken from an 18 month-old female suffering from intractable seizures. Treatment included a hemispherectomy. HCN-1A are frequently used to model neuronal processes for scientific research. HCN-1A cells have been shown to test positive for somatostatin, glutamate, neuron specific enolase, tubulin, vimentin, gamma aminobutyric acid, vasoactive intestinal peptide, and cholecystokinin-8. The cells test negative for glial fibrillary acidic protein, and myelin basis protein. Differentiation of HCN-1A can be induced through culturing the cortical neuronal cells with a mixture of nerve growth factor, dibutyryl cyclic adenosine monophosphate, and 1-isobutyl-3-methylxanthine. When HCN-1A cells differentiate, they take on mature morphology which significantly slows their growth rate, with a doubling time exceeding 120 hours. After HIV/N1T infection, HCN-1A diminished in size and small neurites were observed.
Figure 1. GAPD mRNA levels were quantified using real-time RT-PCR in the HCN1A cells transfected with siRNAs targeting GAPD or non-silencing siRNA. Forty-eight hours post-transfection, the cells were harvested and analyzed by real-time RT-PCR for GAPD mRNA expression levels. Data were normalized against the 18S rRNA signal. Control samples were either mock-transfected or untreated. Values are normalized to untreated sample. Data are means ± SD (n=5).
Figure 2. Protein expression of GAPDH in HCN-1A cells. DNA plasmid expressing GAPDH or siRNA targeting GAPDH were transfected into HCN-1A cells following Altogen Biosystems transfection protocol. At 72 hours post-transfection the cells were analyzed by Western Blot for protein expression levels (normalized by total protein, 10 µg of total protein loaded per each well). Untreated cells used as a negative control.
Selected HCN1A transfection kit citations:
- Journal of Biological Chemistry. 2011. Regulatory Factor X1-induced Down-regulation of Transforming … Feng et al [PDF]
- Cancer Research. 2011 71(15):5144-53. Inhibition of miR-193a expression by… Iliopoulos et al [PDF]
- RNA. 2010 16(11):2108-19. RNase L releases a small RNA from HCV RNA that refolds … Malathi et al [PDF]
Altogen Biosystems transfection and electroporation products for life sciences and cancer research. Company developed two types of in vivo delivery kits for animal research: Tissue-targeted reagents (delivery to liver, pancreas, and kidney tissues), and broad range in vivo delivery reagents (PEG-Liposome, Nanoparticle-based, Lipid-based, and Polymer-based kits). Advanced formulation of reagents and optimized transfection protocols provide efficient intracellular delivery of proteins, DNA, mRNA, shRNA, siRNA, and other negatively charged molecules in vitro and in vivo. Read more about transfection technology at Altogen’s Transfection Resource.
Altogen Labs Research Services:
Altogen Labs provides GLP-compliant contract research studies for pre-clinical research, IND applications, and drug development. Biology CRO services include: Xenograft models (30+), development of stable cell lines, ELISA assay development, cell-based and tissue targeted RNAi studies, safety pharm/tox assays, and other studies (visit AltogenLabs.com).
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