title: Enhancement and optimization of plasmid expression in femtosecond optical transfection.
abstract: Cell transfection using femtosecond lasers is gaining importance for its proven ability to achieve selective transfection in a sterile and relatively non-invasive manner. However, the net efficiency of this technique is limited due to a number of factors that ultimately makes it difficult to be used as a viable and widely used technique. We report here a method to achieve significant enhancement in the efficiency of femtosecond optical transfection. The transfection procedure is modified by incorporating a suitable synthetic peptide containing nuclear localization and DNA binding sequences, assisting DNA import into the nucleus. We achieved a 3-fold enhancement in the transfection efficiency for adherent Chinese Hamster Ovary (CHO-K1) cells with this modified protocol. Further, in the presence of this biochemical reagent, we were able to reduce the required plasmid concentration by ~70% without compromising the transfection efficiency. Also, we report for the first time the successful photo-transfection of recently trypsinised cells with significantly high transfection efficiency when transfected with modified plasmid. This paves the way for the development of high throughput microfluidic optical transfection devices.
authors: SUPA, School of Physics & Astronomy, University of St Andrews, North Haugh, St Andrews, KY16 9SS, Scotland, UK. bb295@st-andrews.ac.uk
authors: Bavishna B
authors: BB
authors: Praveen
authors: David J
authors: DJ
authors: Stevenson
authors: Maciej
authors: M
authors: Antkowiak
authors: Kishan
authors: K
authors: Dholakia
authors: Frank J
authors: FJ
authors: Gunn-Moore
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D055096:Optical Devices; D010957:Plasmids / Q000235:genetics / Q000378:metabolism*; D014025:Titanium / Q000737:chemistry; D014162:Transfection / Q000295:instrumentation / Q000379:methods*
chemical_list: D004247:DNA; D014025:Titanium