Flow cytometry and live confocal analysis for the evaluation of the uptake and intracellular distribution of FITC-ODN into HaCaT cells

Ruozi, Barbara; Montanari, Monica; Vighi, Eleonora; Tosi, Giovanni; Tombesi, Andrea; Battini, Renata; Restani, Cinzia; Leo, Eliana Grazia; Forni, Flavio; Vandelli, Maria Angela

doi:10.1080/08982100902788416

Cited by 6 publications

(6 citation statements)

References 42 publications

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“…In contrast, SP/siRNA complexes did not significantly colocalize with LysoTracker Green (Figure ). This revealed that SP/siRNA complexes were more capable of escaping from the acidic and nuclease rich environment of the lysosomes than LF/siRNA complexes …”

Section: Resultsmentioning

confidence: 99%

“…The intracellular localization of SP or LF/Cy3-siRNA (red) complexes was studied using confocal microscopy. The cells were treated with LysoTracker Green (green), a compound that primarily accumulates in cellular compartments with low internal pH, 36 and siRNA complexes. Confocal microscopy images clearly showed extensive colocalization (yellow) of LF/siRNA complexes and LysoTracker Green.…”

Section: Resultsmentioning

confidence: 99%

“…This revealed that SP/siRNA complexes were more capable of escaping from the acidic and nuclease rich environment of the lysosomes than LF/siRNA complexes. 36 3.7. Intracellular Trafficking Mechanism of the siRNA.…”

Section: Resultsmentioning

confidence: 99%

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SPANosomes as Delivery Vehicles for Small Interfering RNA (siRNA)

et al. 2011

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Non-ionic surfactant vesicles, or SPANosomes (SPs), comprised of cationic lipid and sorbitan monooleate (Span 80) were synthesized and evaluated as siRNA vectors. The SPs had a mean diameter of less than 100 nm and exhibited excellent colloidal stability. The SP/siRNA complexes possessed a slightly positive zeta potential of 12 mV and demonstrated a high siRNA incorporation efficiency of greater than 80%. Cryogenic transmission electron microscopy (cryo-TEM) imaging of the SP/siRNA indicated a predominantly core-shell structure. The SP/siRNA complexes were shown to efficiently and specifically silence expression of both green fluorescent protein (GFP) (66% knockdown) and aromatase (77% knockdown) genes in breast cancer cell lines. In addition, the cellular trafficking pathway of the SP/siRNA was investigated by confocal microscopy using molecular beacons as probes for cytosolic delivery. The results showed efficient endosomal escape and cytosolic delivery of the siRNA cargo following internalization of the SP/siRNA complexes. In conclusion, Span 80 is a potent helper lipid and the SPs are promising vehicles for siRNA delivery.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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SPANosomes as Delivery Vehicles for Small Interfering RNA (siRNA)

et al. 2011

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“…Previously utilised methods of quantitative analysis led either to the destruction of studied objects (i.e. Western blot, Preston, 2002) or left no opportunity to relate the population data to the image of observed object (flow cytometry, Ruozi et al, 2009). The possibility of quantitative analysis of large populations of cells, linked directly with high-resolution imaging is particularly useful for analysing complex tissues such as gut mucosa.…”

Section: Discussionmentioning

confidence: 98%

Electron microscopy finder-grids link quantitative cytometry with high-resolution confocal imaging in the study of TGF-β1 expression in the small intestine mucosa in growing pigs

et al. 2010

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“…Liposomal formulations, prepared with DOTAP (cationic liposomes, sample A), DOTAP:Chol-PEG 3350 -NH 2 1: 0.1 mol:mol (cationic PEGylated liposomes, sample B) were prepared by a thin layer evaporation method followed by extrusion through a 200 nm polycarbonate filter, as previously described [22]. Briefly, lipids (5 mg) were dissolved in 2 ml of chloroform.…”

Section: Cationic and Cationic Pegylated Liposomesmentioning

confidence: 99%