Combined multiphoton imaging-pixel analysis for semiquantitation of skin penetration of gold nanoparticles

Labouta, Hagar I.; Kraus, Tobias; El-Khordagui, Labiba K.; Schneider, Marc

doi:10.1016/j.ijpharm.2011.03.067

Cited by 48 publications

(66 citation statements)

References 14 publications

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“…This qualitative observation with CLSM was quantified by analyzing the weighted number of nanoparticles in the cells by pixel analysis of the CLSM images as described by Labouta et al [35]. The advantage of this method, over manual counting of the number of fluorescent spots, is that it avoids human errors and also considers the area of the fluorescent spots which are larger than the resolution limit.…”

Section: Resultsmentioning

confidence: 99%

“…Images corresponding to a total of 100 cells were acquired for each inhibitor and quantification of number of cells positive for nanoparticle uptake (nanoparticles inside the cells) was carried out by a combined multiphoton-pixel analysis method reported by Labouta et al [35]. This assumption should be valid as long as the coverage of the polymer chain is far from complete.…”

Section: Clsmmentioning

confidence: 99%

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Cellular delivery of polynucleotides by cationic cyclodextrin polyrotaxanes

Dandekar

Jain

Keil

et al. 2012

Journal of Controlled Release

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Cationic polyrotaxanes, obtained by temperature activated threading of cationic cyclodextrin derivatives onto water soluble cationic polymers (ionenes), form metastable nanometric polyplexes with pDNA and combinations of siRNA with pDNA.Because of their low toxicity, the polyrotaxane polyplexes constitute a very interesting system for the transfection of polynucleotides into mammalian cells. The complexation of Cy3-labeled siRNA within the polyplexes was demonstrated by fluorescence correlation spectroscopy. The uptake of the polyplexes (red) was imaged by confocal fluorescence microscopy using the A549 cell line as a model (blue: nuclei, green: membranes). The results prove the potential of polyrotaxanes for further investigations involving knocking down genes of therapeutic interest.

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Section: Resultsmentioning

confidence: 99%

Section: Clsmmentioning

confidence: 99%

Cellular delivery of polynucleotides by cationic cyclodextrin polyrotaxanes

Dandekar

Jain

Keil

et al. 2012

Journal of Controlled Release

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“…For each captured z-stack, optical layers incorporating cell-internalized liposomes were chosen and a z-projection image of the red channel (red fluorescence due to liposomes) was developed using ImageJ. The number of pixels was computed and converted into weighted number of liposomes as described previously [16,17] and as briefly referred to in the methodology of the adhesion experiments.…”

Section: Relative Liposomal Uptakementioning

confidence: 99%

Bacteriomimetic invasin-functionalized nanocarriers for intracellular delivery

Menina

Kochut

Gordon

et al. 2015

Journal of Controlled Release

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Intracellular bacteria invade mammalian cells to establish an infectious niche. The current work models adhesion and subsequent internalization strategy of pathogenic bacteria into mammalian cells to design a bacteriomimetic bioinvasive delivery system. We report on the surface functionalization of liposomes with a C-terminal fragment of invasin (InvA497), an invasion factor in the outer membrane of Yersinia pseudotuberculosis. InvA497-functionalized liposomes adhere to mammalian epithelial HEp-2 cell line at different infection stages with a significantly higher efficiency than liposomes functionalized with bovine serum albumin. Covalent attachment of InvA497 results in higher cellular adhesion than liposomes with physically adsorbed InvA497 with non-specific surface protein alignment. Uptake studies in HEp-2 cells indicate active internalization of InvA497-functionalized liposomes via β1-integrin receptor-mediated uptake mechanism mimicking the natural invasion strategy of Yersinia pseudotuberculosis. Uptake studies in Caco-2 cells at different polarization states demonstrate specific targeting of the InvA497-functionalized liposomes to less polarized cells reflecting the status of inflamed cells. Moreover, when loaded with the anti-infective agent gentamicin and applied to HEp-2 cells infected with Yersinia pseudotuberculosis, InvA497-functionalized liposomes are able to significantly reduce the infection load relative to nonfunctionalized drug-loaded liposomes. This indicates a promising application of such a bacteriomimetic system for drug delivery to intracellular compartments. *Graphical AbstractBacterial invas on into a human cellvia outer ..•

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“…Following fast freezing of the skin specimen, an 8-mm punch in the center of the specimen was placed in a perpendicular direction to the cutting blade to avoid dislocation of the particles from outside into the tissue or vice versa, thus limiting sectioning artifacts. 28 Skin sections were placed on microscopy slides and mounted by an aqueous mounting medium (FluorSaveTM reagent, Calbiochem, San Diego) and covered with glass cover slips.…”

Section: Longitudinal Skin Cryo-sectioningmentioning

confidence: 99%

“…This technique is referred to as multiphoton-absorption-induced luminescence (MAIL), where the absorption of multiple photons from a near-infrared pulsed femtosecond laser can lead to robust luminescence of metal nanoparticles. 8,[23][24][25][26] Multiphoton microscopy was therefore employed as a useful optical technique to track particle penetration in skin 8,22,[26][27][28] and was the imaging tool in our study to track model AuNP.…”

Section: Introductionmentioning

confidence: 99%

Setup for investigating gold nanoparticle penetration through reconstructed skin and comparison to published human skin data

Thude

Schneider

2012

J. Biomed. Opt

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This is an by copyright after embargo allowed publisher's PDF of an article published in Labouta, H.I., Thude, S., Schneider, M. Setup for investigating gold nanoparticle penetration through reconstructed skin and comparison to published human skin data (2013) Journal of Biomedical Optics, 18 (6), art. no. 61218.Setup for investigating gold nanoparticle penetration through reconstructed skin and comparison to published human skin data Abstract. Owing to the limited source of human skin (HS) and the ethical restrictions of using animals in experiments, in vitro skin equivalents are a possible alternative for conducting particle penetration experiments. The conditions for conducting penetration experiments with model particles, 15-nm gold nanoparticles (AuNP), through nonsealed skin equivalents are described for the first time. These conditions include experimental setup, sterility conditions, effective applied dose determination, skin sectioning, and skin integrity check. Penetration at different exposure times (two and 24 h) and after tissue fixation (fixed versus unfixed skin) are examined to establish a benchmark in comparison to HS in an attempt to get similar results to HS experiments presented earlier. Multiphoton microscopy is used to detect gold luminescence in skin sections. λ ex ¼ 800 nm is used for excitation of AuNP and skin samples, allowing us to determine a relative index for particle penetration. Despite the observed overpredictability of penetration into skin equivalents, they could serve as a first fast screen for testing the behavior of nanoparticles and extrapolate their penetration behavior into HS. Further investigations are required to test a wide range of particles of different physicochemical properties to validate the skin equivalent-human skin particle penetration relationship.

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Combined multiphoton imaging-pixel analysis for semiquantitation of skin penetration of gold nanoparticles

Cited by 48 publications

References 14 publications

Cellular delivery of polynucleotides by cationic cyclodextrin polyrotaxanes

Cellular delivery of polynucleotides by cationic cyclodextrin polyrotaxanes

Bacteriomimetic invasin-functionalized nanocarriers for intracellular delivery

Setup for investigating gold nanoparticle penetration through reconstructed skin and comparison to published human skin data

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