Investigating Lyophilization of Lipid Nanocapsules with Fluorescence Correlation Spectroscopy

Smith, Polina B.; Dendramis, Kimberly A.; Chiu, Daniel T.

doi:10.1021/la1003808

Cited by 7 publications

(4 citation statements)

References 24 publications

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“…7,9 However, we recently overcame the stor-age issue for lipid nanocapsules by using an optimized lyophilization procedure that allows us to store lipid nanocapsules over long periods of time at Ϫ20 °C as a lyophilized powder. 19 As a result, we returned to developing light-addressable lipid nanocapsules as a general caging platform for bioactive molecules.…”

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“…Unfortunately, the lipid vesicles were fragile and could not be stored in solution for more than a few days. This practical constraint prompted us to explore a range of more robust nonlipid-based nanocapsules, including ones formed by colloidal templating in silica and polymer-based nanocapsules. , However, we recently overcame the storage issue for lipid nanocapsules by using an optimized lyophilization procedure that allows us to store lipid nanocapsules over long periods of time at −20 °C as a lyophilized powder . As a result, we returned to developing light-addressable lipid nanocapsules as a general caging platform for bioactive molecules.…”

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Intracellular Delivery of Bioactive Molecules using Light-Addressable Nanocapsules

et al. 2010

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This paper describes a method by which molecules that are impermeable to cells are encapsulated in dye-sensitized lipid nanocapsules for delivery into cells via endocytosis. Once inside the cells, the molecules are released from the lipid nanocapsules into the cytoplasm with a single nanosecond pulse from a laser in the far red (645nm). We demonstrate this method with the intracellular release of the second messenger IP 3 in CHO-M1 cells, and report that calcium responses from the cells changed from a sustained increase to a transient spike when the average number of IP 3 released is decreased below 50 molecules per nanocapsule. We also demonstrate the delivery of a 23 kDa AGT fusion protein into Ba/F3 cells to inhibit a key player BCR-ABL in the apoptotic pathway. We show that an average of ~ 8 molecules of the inhibitor is sufficient to induce apoptosis in the majority of Ba/F3 cells. KeywordsLipids; Dye-Sensitized; Nanocapsule; Intracellular; Photolysis To uncover the spatiotemporal dynamics of cellular function, we need techniques that allow us to perturb cells over space and time in a controlled fashion. The most common approach for the delivery of bioactive molecules to cells in a highly spatiotemporally-resolved fashion is the two-photon release of a chemically-caged compound. 1-5 Caged molecules, however, suffer from a number of drawbacks. The design and synthesis of a new caged compound can be tedious and the caging of large protein molecules can be difficult, if not impossible. For intracellular delivery, the caged molecules must be permeable to the cell; this requirement, in turn, makes it difficult to control the precise concentration of the caged molecules inside the cell because of their preferential partitioning into different cellular organelles and membranes. These drawbacks of caged compounds have prompted us to develop lightaddressable lipid nanocapsules as a general platform for caging a wide range of bioactive molecules. 6-12 Nanocapsules based on lipid vesicles represent an emerging class of physical cages,, some of which also have been used recently for intracellular release. [13][14][15][16][17][18] Our first nanocapsules were lipid vesicles which we manipulated with optical tweezers and photolyzed using a single nanosecond laser pulse in the UV. 6,8,10 This capability was sufficient for delivering molecules extracellularly in a cell-culture setting. Unfortunately, the lipid vesicles were fragile and could not be stored in solution for more than a few days. This * To whom correspondence should be addressed chiu@chem.washington.edu. Supporting Information

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Intracellular Delivery of Bioactive Molecules using Light-Addressable Nanocapsules

et al. 2010

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“…Our group has been interested in developing techniques that are capable of probing the structure of biological signaling networks with high spatial and temporal resolution. Towards this end, we have developed a method that employs nanocapsules as physical cages to entrap bioactive molecules within the capsules until the capsules are burst open using a single nanosecond laser pulse 1–8 . Our strategy for using nanocapsules in an in vitro cell culture setting is shown in figure 1.…”

Section: Introduction: Single-site Delivery Of Biological Stimuli mentioning

confidence: 99%

“…We have demonstrated that this method can release the contents of a nanocapsule onto cells with sub-micron spatial resolution and sub-microsecond temporal resolution 1–8 . We have also delivered a wide range of bioactive stimuli to cells, including small molecules and large protein biomolecules.…”

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confidence: 99%

Optical trapping enabled parallel delivery of biological stimuli with high spatial and temporal resolution

2010

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We have developed a method that employs nanocapsules, optical trapping, and single-pulse laser photolysis for delivering bioactive molecules to cells with both high spatial and temporal resolutions. This method is particularly suitable for a cell-culture setting, in which a single nanocapsule can be optically trapped and positioned at a pre-defined location next to the cell, followed by single-pulse laser photolysis to release the contents of the nanocapsule onto the cell. To parallelize this method such that a large array of nanocapsules can be manipulated, positioned, and photolyzed simultaneously, we have turned to the use of spatial light modulators and holographic beam shaping techniques. This paper outlines the progress we have made so far and details the issues we had to address in order to achieve efficient parallel optical manipulations of nanocapsules and particles.

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Nanocapsules-based antimicrobial systems for biomedical applications

Ayshwarya¹,

Prabhu²,

Pichumani³

2023

Antimicrobial Nanosystems

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Investigating Lyophilization of Lipid Nanocapsules with Fluorescence Correlation Spectroscopy

Cited by 7 publications

References 24 publications

Intracellular Delivery of Bioactive Molecules using Light-Addressable Nanocapsules

Intracellular Delivery of Bioactive Molecules using Light-Addressable Nanocapsules

Optical trapping enabled parallel delivery of biological stimuli with high spatial and temporal resolution

Nanocapsules-based antimicrobial systems for biomedical applications

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