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DELIVERY OF MOLECULES INTO CORNEAL ENDOTHELIUM USING NANOPARTICLESACTIVATED BY FEMTOSECOND LASER PULSES: PROOF OF CONCEPTC. Jumelle 1 , N. Campolmi 1 , A. Bernard 1 , S. Piselli 1 , C. Mauclair 2 , E. Audouard 2 , J. Granier 3 , H. Soder 3 , P. Gain 1 , G. Thuret 1¹Corneal Graft Biology, Engineering and Imaging Laboratory, EA2521, Federative Institute of Research in Sciences andHealth Engineering, Faculty of Medicine, Jean Monnet University; ²Laboratoire Hubert Curien; ³IMPULSION SAS, PôleOptique Vision, FrancePurpose: The so-called NanoFemtoTransfection (NFT) is an innovative and promising non-viraltechnique to transfer molecules into cells (Charatasky. Nature nanotechnology 2011). It consists intemporarily permeabilizing cell membrane by a photoacoustic effect obtained by nanoparticles ofblack carbon activated by Ti-Saphir femtosecond laser (fsL) pulses. Calcein (622 Da), tagged bovineserum albumine (70 kDa) and one eGFP plasmid (5 MDa) were transfected into two non-adherent celllines (DU145 prostate-cancer and GS-9L rat gliosarcoma). Our aim was to adapt the NFT to adherenthuman corneal endothelial cells (HCEC).Methods: We tested the NFT of calcein in vitro on the HCEC line HCEC-12 seeded at 1500cells/mm2 in 6 wells plates and ex vivo on whole human organ cultured corneas. A matrix of experimentscomprising 4 exposition times, 6 fluences and 2 fsL beam movements was performed in order toobtain transfection with minimal toxicity. After exposition to fsL, nuclei were counterstained withHoechst33342 and transfection efficiency was determined by observation on a fluorescence invertedmicroscope (IX81, Olympus, Japan) and further quantified by flow cytometry (FACSCalibur, BD, CA).Viability was assessed by Trypan blue staining.Results: In HCEC-12, a fluence of 100 mJ/cm² and a laser beam movement of 3,5 mm/s gave atransfection of 17% and a viability of 97%. In whole corneas, with the same parameters, transfectionwas detectable in disseminated EC.Conclusions: We obtained the POC of the NFT in HCEC. Further optimization is ongoing to increasethe transfection rate while maintaining minimal toxicity, especially for bigger molecules, like plasmids.XXV ANNUAL MEETING OF THE EUROPEAN EYE BANK ASSOCIATION Zagreb, Croatia 18/19 January 2013 55