Evaluation of pulsed laser ablation in liquids generated gold nanoparticles as novel transfection tools: efficiency and cytotoxicity

Varying transfection efficiencies and cytotoxicity are crucial aspects in cell manipulation. The utilization of gold nanoparticles (AuNP) has lately attracted special interest to enhance transfection efficiency. Conventional AuNP are usually generated by chemical reactions or gas pyrolysis requiring often cell-toxic stabilizers or coatings to conserve their characteristics. Alternatively, stabilizer- and coating-free, highly pure, colloidal AuNP can be generated by pulsed laser ablation in liquids (PLAL). Mammalian cells were transfected efficiently by addition of PLAL-AuNP, but data systematically evaluating the cell-toxic potential are lacking. Herein, the transfection efficiency and cytotoxicity of PLAL AuNP was evaluated by transfection of a mammalian cell line with a recombinant HMGB1/GFP DNA expression vector. Different methods were compared using two sizes of PLAL-AuNP, commercialized AuNP, two magnetic NP-based protocols and a conventional transfection reagent (FuGENE HD; FHD). PLAL-AuNP were generated using a Spitfire Pro femtosecond laser system delivering 120 fs laser pulses at a wavelength of 800 nm focusing the fs-laser beam on a 99.99% pure gold target placed in ddH2O. Transfection efficiencies were analyzed after 24h using fluorescence microscopy and flow cytometry. Toxicity was assessed measuring cell proliferation and percentage of necrotic, propidium iodide positive cells (PI %). The addition of PLAL-AuNP significantly enhanced transfection efficiencies (FHD: 31 %; PLAL-AuNP size-1: 46 %; size-2: 50 %) with increased PI% but no reduced cell proliferation. Commercial AuNP-transfection showed significantly lower efficiency (23 %), slightly increased PI % and reduced cell proliferation. Magnetic NP based methods were less effective but showing also lowest cytotoxicity. In conclusion, addition of PLAL-AuNP provides a novel tool for transfection efficiency enhancement with acceptable cytotoxic side-effects.

[1]  Sabine Neuss,et al.  Size-dependent cytotoxicity of gold nanoparticles. , 2007, Small.

[2]  Alexander Heisterkamp,et al.  TNF-α induced secretion of HMGB1 from non-immune canine mammary epithelial cells (MTH53A). , 2012, Cytokine.

[3]  E. Tekle,et al.  Electroporation by using bipolar oscillating electric field: an improved method for DNA transfection of NIH 3T3 cells. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[4]  S. Petersen,et al.  Golden perspective: application of laser-generated gold nanoparticle conjugates in reproductive biology. , 2011, Reproduction in domestic animals = Zuchthygiene.

[5]  C. Murphy,et al.  Gold nanoparticles are taken up by human cells but do not cause acute cytotoxicity. , 2005, Small.

[6]  Stephan Barcikowski,et al.  In Situ Bioconjugation: Single Step Approach to Tailored Nanoparticle‐Bioconjugates by Ultrashort Pulsed Laser Ablation , 2009 .

[7]  Jennifer A. Dahl,et al.  Toward Greener Nanosynthesis , 2007 .

[8]  Jun Liu,et al.  Antibody and DNA dual-labeled gold nanoparticles: Stability and reactivity , 2008 .

[9]  E. Papapetrou,et al.  Genetic modification of hematopoietic stem cells with nonviral systems: past progress and future prospects , 2005, Gene Therapy.

[10]  Kevin D Bunting,et al.  Safety concerns related to hematopoietic stem cell gene transfer using retroviral vectors. , 2004, Current gene therapy.

[11]  Pablo Menendez,et al.  Human embryonic stem cells , 2007, Stem Cell Reviews.

[12]  T. Kondow,et al.  Dissociation and Aggregation of Gold Nanoparticles under Laser Irradiation , 2001 .

[13]  Yoshihiro Takeda,et al.  Full Physical Preparation of Size-Selected Gold Nanoparticles in Solution: Laser Ablation and Laser-Induced Size Control , 2002 .

[14]  R. Shukla,et al.  Biocompatibility of gold nanoparticles and their endocytotic fate inside the cellular compartment: a microscopic overview. , 2005, Langmuir : the ACS journal of surfaces and colloids.

[15]  Stephan Barcikowski,et al.  In situ bioconjugation—Novel laser based approach to pure nanoparticle-conjugates , 2009 .

[16]  Monic Shah,et al.  Biological applications of gold nanoparticles. , 2014, Journal of nanoscience and nanotechnology.

[17]  Chad A. Mirkin,et al.  Oligonucleotide-Modified Gold Nanoparticles for Intracellular Gene Regulation , 2006, Science.

[18]  J. Northrop,et al.  Lipofection: a highly efficient, lipid-mediated DNA-transfection procedure. , 1987, Proceedings of the National Academy of Sciences of the United States of America.

[19]  Stephan Barcikowski,et al.  Co-transfection of plasmid DNA and laser-generated gold nanoparticles does not disturb the bioactivity of GFP-HMGB1 fusion protein , 2009, Journal of nanobiotechnology.

[20]  Vincent M Rotello,et al.  Gold nanoparticles in delivery applications. , 2008, Advanced drug delivery reviews.