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Subject: Protein delivery ×

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    Silica nanoparticles for intracellular protein delivery: A novel synthesis approach using green fluorescent protein
    (London : BioMed Central, 2017) Schmidt, Sarah; Tavernaro, Isabella; Cavelius, Christian; Weber, Eva; Kümper, Alexander; Schmitz, Carmen; Fleddermann, Jana; Kraegeloh, Annette
    In this study, a novel approach for preparation of green fluorescent protein (GFP)-doped silica nanoparticles with a narrow size distribution is presented. GFP was chosen as a model protein due to its autofluorescence. Protein-doped nanoparticles have a high application potential in the field of intracellular protein delivery. In addition, fluorescently labelled particles can be used for bioimaging. The size of these protein-doped nanoparticles was adjusted from 15 to 35 nm using a multistep synthesis process, comprising the particle core synthesis followed by shell regrowth steps. GFP was selectively incorporated into the silica matrix of either the core or the shell or both by a one-pot reaction. The obtained nanoparticles were characterised by determination of particle size, hydrodynamic diameter, ζ-potential, fluorescence and quantum yield. The measurements showed that the fluorescence of GFP was maintained during particle synthesis. Cellular uptake experiments demonstrated that the GFP-doped nanoparticles can be used as stable and effective fluorescent probes. The study reveals the potential of the chosen approach for incorporation of functional biological macromolecules into silica nanoparticles, which opens novel application fields like intracellular protein delivery.
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    Switchable Release of Bone Morphogenetic Protein from Thermoresponsive Poly(NIPAM-co-DMAEMA)/Cellulose Sulfate Particle Coatings
    (Basel : MDPI, 2018) Müller, Martin; Urban, Birgit; Reis, Berthold; Yu, Xiaoqian; Grab, Anna Luise; Cavalcanti-Adam, Elisabetta Ada; Kuckling, Dirk
    Thermoresponsive coatings of poly(N-isopropylacrylamide-co-DMAEMA)/cellulose sulfate (PNIPAM-DMAEMA/CS) complexes are reported eluting bone-morphogenetic-protein-2 (BMP-2) on demand relevant for implant assisted local bone healing. PNIPAM-DMAEMA/CS dispersions contained colloid particles with hydrodynamic radii RH = 170–288 nm at T = 25 °C shrinking to RH = 74–103 nm at T = 60 °C. Obviously, PNIPAM-DMAEMA/CS undergoes volume phase transition (VPT) analogously to pure PNIPAM, when critical VPT temperature (VPTT) is exceeded. Temperature dependent turbidity measurements revealed broad VPT and VPTT 47 °C for PNIPAM-DMAEMA/CS colloid dispersions at pH = 7.0. FTIR spectroscopy on thermoresponsive PNIPAM-DMAEMA/CS particle coatings at germanium model substrates under HEPES buffer indicated both wet-adhesiveness and VPT behavior based on diagnostic band intensity increases with temperature. From respective temperature courses empirical VPTT ≈ 42 °C for PNIPAM-DMAEMA/CS coatings at pH = 7.0 were found, which were comparable to VPTT found for respective dispersions. Finally, the PNIPAM-DMAEMA/CS coatings were loaded with BMP-2 and model protein papain (PAP). Time dependent FTIR spectroscopic measurements showed, that for T = 37 °C there was a relative protein release of ≈30% for PAP and ≈10% for BMP-2 after 24 h, which did not increase further. Heating to T = 42 °C for PAP and to 47 °C for BMP-2 further secondary protein release of ≈20% after 24 h was found, respectively, interesting for clinical applications. BMP-2 eluted even at 47 °C was found to be still biologically active