Non-invasive imaging of cells that have been labeled non-destructively, such as with nanoemulsions or reporter genes that can be detected by molecular imaging, to monitor their location, viability, cell lineage expansion, response to drugs, movement, or other behaviors in vivo.
1Department of Flow and Image Cytometry, Roswell Park Cancer Institute, 2Flow Cytometry & Cell Sorting Resource Laboratory, University of Pennsylvania, 3SciGro, Inc., 4Department of Pathology and Laboratory Medicine, University of Pennsylvania
Successful use of cell tracking dyes to monitor immune cell function and proliferation involves several critical steps. We describe methods for: 1) obtaining bright, uniform, reproducible label-ing with membrane dyes; 2) selecting fluorochromes and data acquisition conditions; and 3) choosing a model to quantify cell proliferation based on dye dilution.
Published December 13, 2012. Keywords: Cellular Biology, Molecular Biology, Cell tracking, PKH26, CFSE, membrane dyes, dye dilution, proliferation modeling, lymphocytes
JoVE Clinical and Translational Medicine
1Department of Tumor Immunology, Nijmegen Center for Molecular Life Sciences, Radboud University Medical Center, 2In-Vivo-NMR Laboratory, Max Planck Institute for Neurological Research, 3German Center for Neurodegenerative Diseases (DZNE)
We describe a general protocol for in vivo cell tracking using MRI in a mouse model with ex vivo labeled cells. A typical protocol for cell labeling, image acquisition processing and quantification is included.
Published November 25, 2013. Keywords: Medicine, Animal Models, Immune System Diseases, MRI, 19F MRI, Cell Tracking, Quantification, Cell Label, In vivo Imaging
1Max Delbrück Center for Molecular Medicine
Here, we present a method for the photoactivated switch of photoconvertible fluorescent proteins (PCFPs) in the living zebrafish embryo and further tracking of photoconverted protein at specific time points during development. This methodology allows monitoring of cell biological events underlying different developmental processes in a live vertebrate organism.
Published September 28, 2012. Keywords: Developmental Biology, Cellular Biology, Molecular Biology, Cell tracking, live cell imaging, photoconvertible fluorescent proteins, tissue morphogenesis, Danio rerio, zebrafish, embryo
1Biomedical Engineering Program, University of South Carolina, 2Chemical Engineering Department, University of South Carolina
A method for photo-encapsulation of cells in a crosslinked PEG hydrogel is described. Hypoxic signaling within encapsulated murine insulinoma (MIN6) aggregates is tracked using a fluorescent marker system. This system allows serial examination of cells within a hydrogel scaffold and correlation of hypoxic signaling with changes in cell phenotype.
Published December 16, 2011. Keywords: Bioengineering, Cell encapsulation, PEG, cell aggregation, hypoxia, insulin secretion, fluorescent imaging
1Department of Ophthalmology, Massachusetts Eye and Ear, 2JoVE Content Production
Published August 1, 2014. Keywords: This Month in JoVE,
1Department of Bionanoscience, Kavli Institute of Nanoscience, Delft University of Technology
Magnetic tweezers, a powerful single-molecule manipulation technique, can be adapted for the direct measurements of the twist (using a configuration called freely-orbiting magnetic tweezers) and torque (using a configuration termed magnetic torque tweezers) in biological macromolecules. Guidelines for performing such measurements are given, including applications to the study of DNA and associated nucleo-protein filaments.
Published May 19, 2014. Keywords: Bioengineering, magnetic tweezers, magnetic torque tweezers, freely-orbiting magnetic tweezers, twist, torque, DNA, single-molecule techniques
1Center for Theoretical Problems of Physicochemical Pharmacology, Russian Academy of Sciences, 2Federal Research Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russia, 3Physiology Department, Perelman School of Medicine, University of Pennsylvania
Microtubules are inherently unstable polymers, and their switching between growth and shortening is stochastic and difficult to control. Here we describe protocols using segmented microtubules with photoablatable stabilizing caps. Depolymerization of segmented microtubules can be triggered with high temporal and spatial resolution, thereby assisting analysis of motions with the disassembling microtubule ends.
Published March 15, 2014. Keywords: Basic Protocol, microscopy flow chamber, single-molecule fluorescence, laser trap, microtubule-binding protein, microtubule-dependent motor, microtubule tip-tracking
JoVE Immunology and Infection
1Microbiology Unit, Department of Biochemistry, University of Oxford, 2Biological Physics Research Group, Clarendon Laboratory, Department of Physics, University of Oxford
Photoactivated localization microscopy (PALM) combined with single-molecule tracking allows direct observation and quantification of protein-DNA interactions in live Escherichia coli cells.
Published March 10, 2014. Keywords: Immunology, Super-resolution microscopy, single-particle tracking, Live-cell imaging, DNA-binding proteins, DNA repair, molecular diffusion
1Department of Biology, Boston College
The MATLAB-based, open source software package, plusTipTracker, can be used to analyze image series of fluorescently-labeled +TIPs to quantify microtubule dynamics.
Published September 7, 2014. Keywords: Molecular Biology, plusTipTracker, microtubule plus-end-tracking proteins, EB1, growth cone, Xenopus laevis, live cell imaging analysis, microtubule dynamics
1Chemical and Biomolecular Engineering Department, University of Houston
Confocal microscopy is used to image quiescent and flowing colloid-polymer mixtures, which are studied as model systems for attractive suspensions. Image analysis algorithms are used to calculate structural and dynamic metrics for the colloidal particles that measure changes due to geometric confinement.
Published May 20, 2014. Keywords: Chemistry, confocal microscopy, particle tracking, colloids, suspensions, confinement, gelation, microfluidics, image correlation, dynamics, suspension flow