Biolistic transfection of organotypic cultures of rat visual cortex using a handheld device.
Our aim is the biolistic transfection of organotypic cultures of rat visual cortex with plasmids coding for neurotrophic factors, which then become expressed for limited periods of time during postnatal ontogenesis. Out of two commercially available devices, we adopted the handheld 'Helios Gene Gun' instead of the stationary PDS-1000He (both Biorad, Munich, Germany). This device allows multiple transfections of single targets and the transfection of a distinct part of a co-culture when utilising an aperture. Unfortunately, the most detailed protocols are limited to the stationary device and not compatible with the hand-held device. We report here the construction of a support for the gene gun including an aperture and the establishment of a protocol to efficiently transfect rat cortical slice cultures. We achieve a high degree of co-expression of independent plasmids coated on the same particles. The expression of the neurotrophin plasmids is demonstrated on mRNA and protein level.
Wirth
Marcus J
MJ
AG Entwicklungsneurobiologie, Fakultät für Biologie, Ruhr-Universität, ND 6/56a, D-44780, Bochum, Germany. marcus.wirth@ruhr-uni-bochum.de
Wahle
Petra
P
D000818:Animals; D000831:Animals, Newborn; D019470:Biolistics / Q000379:methods*; D019208:Brain-Derived Neurotrophic Factor / Q000378:metabolism; D002356:Cartilage / Q000502:physiology; D018920:Coculture Techniques; D015870:Gene Expression / Q000502:physiology*; D017979:Gold Colloid; D049452:Green Fluorescent Proteins; D017403:In Situ Hybridization; D007202:Indicators and Reagents / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D009414:Nerve Growth Factors / Q000378:metabolism; D009474:Neurons / Q000378:metabolism; D009924:Organ Culture Techniques; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D014162:Transfection / Q000379:methods; D024723:Ventricular Myosins / Q000378:metabolism; D014793:Visual Cortex / Q000166:cytology / Q000378:metabolism*; D000097573:Red Fluorescent Protein
D019208:Brain-Derived Neurotrophic Factor; D017979:Gold Colloid; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D009414:Nerve Growth Factors; D012333:RNA, Messenger; D024723:Ventricular Myosins; D000097573:Red Fluorescent Protein