| 1. |
- Enger, Jonas, 1966, et al.
(författare)
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Optical tweezers applied to a microfluidic system
- 2004
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Ingår i: Lab on a Chip. - : Royal Society of Chemistry (RSC). - 1473-0197 .- 1473-0189. ; 4, s. 196-200
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Tidskriftsartikel (refereegranskat)abstract
- We will demonstrate how optical tweezers can be combined with a microfluidic system to create a versatile microlaboratory. Cells are moved between reservoirs filled with different media by means of optical tweezers. We show that the cells, on a timescale of a few seconds, can be moved from one reservoir to another without the media being dragged along with them. The system is demonstrated with an experiment where we expose E. coli bacteria to different fluorescent markers. We will also discuss how the system can be used as an advanced cell sorter. It can favorably be used to sort out a small fraction of cells from a large population, in particular when advanced microscopic techniques are required to distinguish various cells. Patterns of channels and reservoirs were generated in a computer and transferred to a mask using either a sophisticated electron beam technique or a standard laser printer. Lithographic methods were applied to create microchannels in rubber silicon (PDMS). Media were transported in the channels using electroosmotic flow. The optical system consisted of a combined confocal and epi-fluorescence microscope, dual optical tweezers and a laser scalpel.
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| 2. |
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| 3. |
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| 4. |
- Goksör, Mattias, 1975, et al.
(författare)
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Optical manipulation in combination with multiphoton microscopy for single-cell studies
- 2004
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Ingår i: Applied Optics. ; 43:25, s. 4831-4837
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Tidskriftsartikel (refereegranskat)abstract
- We demonstrate how optical tweezers can be incorporated into a multiphoton microscope to achieve three-dimensional imaging of trapped cells. The optical tweezers, formed by a cw 1064 nm Nd:YVO4 laser, were used to trap live yeast cells in suspension while the 4′, 6-diamidino-2-phenylindole-stained nucleus was imaged in three dimensions by use of a pulsed femtosecond laser. The trapped cell was moved in the axial direction by changing the position of an external lens, which was used to control the divergence of the trapping laser beam. This gives us a simple method to use optical tweezers in the laser scanning of confocal and multiphoton microscopes. It is further shown that the same femtosecond laser as used for the multiphoton imaging could also be used as laser scissors, allowing us to drill holes in the membrane of trapped spermatozoa.
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| 5. |
- Persson, Daniel, 1972, et al.
(författare)
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Vesicle size-dependent translocation of penetratin analogs across lipid membranes
- 2004
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Ingår i: Biochimica et Biophysica Acta - Biomembranes. - : Elsevier BV. - 1879-2642 .- 0005-2736. ; 1665:1-2, s. 142-155
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Tidskriftsartikel (refereegranskat)abstract
- The recent discoveries of serious artifacts associated with the use of cell fixation in studies of the cellular uptake of cell-penetrating peptides (CPPs) have prompted a reevaluation of the current understanding of peptide-mediated cellular delivery. Following a report on the differential cellular uptake of a number of penetratin analogs in unfixed cells, we here investigate their membrane translocation abilities in large and giant unilamellar vesicles (LUVs and GUVs, respectively). Surprisingly, in contrast to the behavior in living cells, all peptides readily entered the giant vesicles ( > 1 μm) as proved by confocal microscopy, while none of them could cross the membranes of LUVs (100 nm). For determination of the location of the peptides in the LUVs, a new concept was introduced, based on sensitive resonance energy transfer (RET) measurements of the enhanced fluorescence of acceptor fluorophores present solely in the inner leaflet. An easily adopted method to prepare such asymmetrically labeled liposomes is described. The membrane insertion depths of the tryptophan moieties of the peptides were determined by use of brominated lipids and found to be very similar for all of the peptides studied. We also demonstrate that infrared spectroscopy on the lipid carbonyl stretch vibration peak is a convenient technique to determine phospholipid concentration. © 2004 Elsevier B.V. All rights reserved.
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| 6. |
- Prikulis, Juris, 1973, et al.
(författare)
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Optical spectroscopy of single trapped metal nanoparticles in solution
- 2004
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Ingår i: Nano letters (Print). - : American Chemical Society (ACS). - 1530-6984 .- 1530-6992. ; 4:1, s. 115-118
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Tidskriftsartikel (refereegranskat)abstract
- We present a simple method for the optical manipulation and spectroscopy of colloidal silver nanoparticles in aqueous solution using optical tweezers combined with dark-field microscopy. Because of their localized plasmon resonances, single trapped metal nanoparticles can be used as efficient near-field optical probes, with potential applicability in surface-enhanced spectroscopy, near-field microscopy, and biochemical sensing schemes. As a proof of principle, we study the near-field optical interaction between a trapped and an immobilized Ag nanoparticle.
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| 7. |
- Ramser, Kerstin, et al.
(författare)
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Resonance Raman spectroscopy of optically trapped functional erythrocytes
- 2004
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Ingår i: Journal of Biomedical Optics. - : SPIE-Intl Soc Optical Eng. - 1083-3668 .- 1560-2281. ; 9:3, s. 593-600
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Tidskriftsartikel (refereegranskat)abstract
- We introduce a novel setup combining a micro-Raman spectrometer with external optical tweezers, suitable for resonance Raman studies of single functional trapped cells. The system differs from earlier setups in that two separate laser beams used for trapping and Raman excitation are combined in a double-microscope configuration. This has the advantage that the wavelength and power of the trapping and probe beam can be adjusted individually to optimize the functionality of the setup and to enable the recording of resonance Raman profiles from a single trapped cell. Trapping is achieved by tightly focusing infrared (IR) diode laser radiation (830 nm) through an inverted oil-immersion objective, and resonance Raman scattering is excited by the lines of an argon:krypton ion laser. The functionality of the system is demonstrated by measurements of trapped single functional erythrocytes using different excitation lines (488.0, 514.5, and 568.2 nm) in resonance with the heme moiety and by studying spectral evolution during illumination. We found that great care has to be taken in order to avoid photodamage caused by the visible Raman excitation, whereas the IR trapping irradiation does not seem to harm the cells or alter the hemoglobin Raman spectra. Stronger photodamage is induced by Raman excitation using 488.0- and 514.5-nm irradiation, compared with excitation with the 568.2-nm line.
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| 9. |
- Thoren, Per, 1972, et al.
(författare)
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Membrane binding and translocation of cell-penetrating peptides
- 2004
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 1520-4995 .- 0006-2960. ; 43:12, s. 3471-3489
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Tidskriftsartikel (refereegranskat)abstract
- Cell-penetrating peptides (CPPs) have been extensively studied during the past decade, because of their ability to promote the cellular uptake of various cargo molecules, e.g., oligonucleotides and proteins. In a recent study of the uptake of several analogues of penetratin, Tat(48-60) and oligoarginine in live (unfixed) cells [Thorén et al. (2003) Biochem. Biophys. Res. Commun. 307, 100-107], it was found that both endocytotic and nonendocytotic uptake pathways are involved in the internalization of these CPPs. In the present study, the membrane interactions of some of these novel peptides, all containing a tryptophan residue to facilitate spectroscopic studies, are investigated. The peptides exhibit a strong affinity for large unilamellar vesicles (LUVs) containing zwitterionic and anionic lipids, with binding constants decreasing in the order penetratin > R 7 W > TatP59W > TatLysP59W. Quenching studies using the aqueous quencher acrylamide and brominated lipids indicate that the tryptophan residues of the peptides are buried to a similar extent into the membrane, with an average insertion depth of ∼10-11 Å from the bilayer center. The membrane topology of the peptides was investigated using an assay based on resonance energy transfer between tryptophan and a fluorescently labeled lysophospholipid, lysoMC, distributed asymmetrically in the membranes of LUVs. By determination of the energy transfer efficiency when peptide was added to vesicles with lysoMC present exclusively in the inner leaflet, it was shown that none of the peptides investigated is able to translocate across the lipid membranes of LUVs. By contrast, confocal laser scanning microscopy studies on carboxyfluorescein-labeled peptides showed that all of the peptides rapidly traverse the membranes of giant unilamellar vesicles (GUVs). The choice of model system is thus crucial for the conclusions about the ability of CPPs to translocate across lipid membranes. Under the conditions used in the present study, peptide-lipid interactions alone cannot explain the different cellular uptake characteristics exhibited by these peptides.
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