| 1. |
- Forslund, E., et al.
(författare)
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Novel microchip-based tools facilitating live cell imaging and assessment of functional heterogeneity within NK cell populations
- 2012
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Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 3:OCT, s. 300-
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Tidskriftsartikel (refereegranskat)abstract
- Each individual has a heterogeneous pool of NK cells consisting of cells that may be specialized towards specific functional responses such as secretion of cytokines or killing of tumor cells. Many conventional methods are not fit to characterize heterogeneous populations as they measure the average response of all cells. Thus, there is a need for experimental platforms that provide single cell resolution. In addition, there are transient and stochastic variations in functional responses at the single cell level, calling for methods that allow studies of many events over extended periods of time. This paper presents a versatile microchip platform enabling long-term microscopic studies of individual NK cells interacting with target cells. Each microchip contains an array of microwells, optimized for medium or high-resolution time-lapse imaging of single or multiple NK and target cells, or for screening of thousands of isolated NK-target cell interactions. Individual NK cells confined with target cells in small microwells is a suitable setup for high-content screening and rapid assessment of heterogeneity within populations, while microwells of larger dimensions are appropriate for studies of NK cell migration and sequential interactions with multiple target cells. By combining the chip technology with ultrasonic manipulation, NK and target cells can be forced to interact and positioned with high spatial accuracy within individual microwells.This setup effectively and synchronously creates NK-target conjugates at hundreds of parallel positions in the microchip. Thus, this facilitates assessment of temporal aspects of NK-target cell interactions, e.g., conjugation, immune synapse formation, and cytotoxic events.The microchip platform presented here can be used to effectively address questions related to fundamental functions of NK cells that can lead to better understanding of how the behavior of individual cells add up to give a functional response at the population level.
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| 2. |
- Bertilson, Michael, et al.
(författare)
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Laboratory soft-x-ray microscope for cryotomography of biological specimens
- 2011
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Ingår i: Optics Letters. - 0146-9592 .- 1539-4794. ; 36:14, s. 2728-2730
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Tidskriftsartikel (refereegranskat)abstract
- Soft-x-ray cryotomography allows quantitative and high-resolution three-dimensional imaging of intact unstained cells. To date, the method relies on synchrotron-radiation sources, which limits accessibility for researchers. Here we present a laboratory water-window microscope for cryotomography. It is based on a lambda = 2.48nm liquid-jet laser-plasma source, a normal-incidence multilayer condenser, a 30nm zone-plate objective, and a cryotilt sample holder. We demonstrate high-resolution imaging, as well as quantitative tomographic imaging, of frozen intact cells. The reconstructed tomogram of the intracellular local absorption coefficient shows details down to similar to 100nm.
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| 3. |
- Christakou, Athanasia. E., et al.
(författare)
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Aggregation and long-term positioning of cells by ultrasound in a multi-well microchip for high-resolution imaging of the natural killer cell immune synapse
- 2011
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Ingår i: 15th International Conference on Miniaturized Systems for Chemistry and Life Sciences 2011, MicroTAS 2011. - 9781618395955 ; , s. 329-331
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Konferensbidrag (refereegranskat)abstract
- In this study we investigate the ability of Natural Killer (NK) cells to form ultrasound-mediated intercellular contacts with target cells in a multi-well microdevice by high-resolution confocal-microscopy imaging of inhibitory immune synapses. Furthermore, we compare the NK-Target cell cluster migration with and without ultrasound actuation. Experiments indicate that clusters of cells are positioned and maintained centered in the wells for 17 hours when they are exposed continuously to ultrasound. Our system can be used for both screening high numbers of events in low resolution and also for high resolution imaging of long term cell-cell interactions.
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| 4. |
- Christakou, Athanasia E., et al.
(författare)
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Characterization of natural killer cells' cytotoxic heterogeneity using an array of sono-cages
- 2012
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Ingår i: Proceedings of the 16th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2012. - : Chemical and Biological Microsystems Society. - 9780979806452 ; , s. 1555-1557
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Konferensbidrag (refereegranskat)abstract
- Using a multi-well device as an array of sono-cages for single cell analysis, we quantify for the first time the heterogeneity of natural killer (NK) cells' cytotoxic response against cancer cells. We report a fraction of inactive NK cells within the tested population (36%), as well as the existence of few 'serial killers' that eliminate up to six targets during 4 hours. We also characterize the multi-well acoustic device in terms of trapping efficiency at different actuation voltages, using adherent and non-adherent cell lines. We show that the acoustic forces applied on the cells can be compared to forces of biological processes (i.e. cell adherence).
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| 5. |
- Christakou, Athanasia E., et al.
(författare)
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Live cell imaging in a micro-array of acoustic traps facilitates quantification of natural killer cell heterogeneity
- 2013
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Ingår i: Integrative Biology. - : Oxford University Press (OUP). - 1757-9694 .- 1757-9708. ; 5:4, s. 712-719
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Tidskriftsartikel (refereegranskat)abstract
- Natural killer (NK) cells kill virus-infected or cancer cells through the release of cytotoxic granules into a tight intercellular contact. NK cell populations comprise individual cells with varying sensitivity to distinct input signals, leading to disparate responses. To resolve this NK cell heterogeneity, we have designed a novel assay based on ultrasound-assisted cell-cell aggregation in a multiwell chip allowing high-resolution time-lapse imaging of one hundred NK-target cell interactions in parallel. Studying human NK cells' ability to kill MHC class I deficient tumor cells, we show that approximately two thirds of the NK cells display cytotoxicity, with some NK cells being particularly active, killing up to six target cells during the assay. We also report that simultaneous interaction with several susceptible target cells increases the cytotoxic responsiveness of NK cells, which could be coupled to a previously unknown regulatory mechanism with implications for NK-mediated tumor elimination.
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| 6. |
- Christakou, Athanasia E., et al.
(författare)
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Solid tumor spheroid formation by temperature-controlled high voltage ultrasound in a multi-well microdevice
- 2014
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Ingår i: 18th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2014. - : Chemical and Biological Microsystems SocietyChemical and Biological Microsystems Society. - 9780979806476 ; , s. 573-575
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Konferensbidrag (refereegranskat)abstract
- In the present work we demonstrate effective 3D growth of human hepatocellular carcinoma (HCC) HepG2 cell spheroids in parallel in a multi-well microdevice actuated with high voltage ultrasound in a temperature-controlled system. We compare the spheroid formation during continuous ultrasound exposure for one week where we formed spheroids in 59% of the wells, with the spheroid formation without ultrasound actuation, where we obtained 0% spheroids. Furthermore, we present an application of the tumor spheroids for investigating natural killer (NK) cells behavior against solid tumors.
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| 7. |
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| 8. |
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| 9. |
- Fogelqvist, Emelie, et al.
(författare)
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The Stockholm laboratory cryo x-ray microscope : towards cell-cell interaction studies
- 2013
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Ingår i: 11th International Conference On X-Ray Microscopy (XRM2012). - : Institute of Physics (IOP). ; , s. 012054-
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Konferensbidrag (refereegranskat)abstract
- We describe recent improvements in the Stockholm laboratory x-ray microscope and the first experiments aiming towards studies of cell-cell interaction. The shorter exposure time due to a higher brightness laser-plasma source will become of large importance for tomography while the reproducible cryo preparation of few-cell samples is essential for the interaction studies.
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| 10. |
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