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University of Utah

6 ARTICLES PUBLISHED IN JoVE

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Biology

Determination of Mammalian Cell Counts, Cell Size and Cell Health Using the Moxi Z Mini Automated Cell Counter
Gregory M. Dittami 1, Manju Sethi 1, Richard D. Rabbitt 2, H. Edward Ayliffe 1
1Orflo Technologies, 2University of Utah

The Moxi Z miniature automated cell counter is a novel instrument that combines the Coulter Principle with patented thin-film sensor technology and a proprietary software algorithm to perform sizing and counting of a broad size range of particles as well as to determine the overall health of monodisperse mammalian cell cultures. This protocol describes the use of this instrument for counting and assessing the health of cell cultures.

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Bioengineering

Hydrophobic Salt-modified Nafion for Enzyme Immobilization and Stabilization
Shannon Meredith 1, Shuai Xu 1, Matthew T. Meredith 1, Shelley D. Minteer 1
1Departments of Chemistry and Materials Science and Engineering, University of Utah

This article will describe the procedure for synthesizing a hydrophobically modified Nafion enzyme immobilization membrane and how to immobilize proteins and/or enzymes within the membrane and test their specific activity.

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Biology

Nano-fEM: Protein Localization Using Photo-activated Localization Microscopy and Electron Microscopy
Shigeki Watanabe 1, Jackson Richards 1, Gunther Hollopeter 1, Robert J. Hobson 1, Wayne M. Davis 1, Erik M. Jorgensen 1
1Department of Biology, Howard Hughes Medical Institute, University of Utah

We describe a method to localize fluorescently tagged proteins in electron micrographs. Fluorescence is first localized using photo-activated localization microscopy on ultrathin sections. These images are then aligned to electron micrographs of the same section.

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Biology

Analysis of Gene Function and Visualization of Cilia-Generated Fluid Flow in Kupffer's Vesicle
Guangliang Wang 1, H. Joseph Yost 2, Jeffrey D. Amack 1
1Department of Cell and Developmental Biology, State University of New York, Upstate Medical University, 2Department of Neurobiology and Anatomy, Eccles Institute of Human Genetics, University of Utah

Cilia-generated fluid flow in Kupffer’s Vesicle (KV) controls left-right patterning of the zebrafish embryo. Here, we describe a technique to modulate gene function specifically in KV cells. In addition, we show how to deliver fluorescent beads into KV to visualize fluid flow.

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Engineering

Construction of a High Resolution Microscope with Conventional and Holographic Optical Trapping Capabilities
Jacqualine Butterfield 1, Weili Hong 1, Leslie Mershon 1, Michael Vershinin 1
1Department of Physics and Astronomy, University of Utah

The system described herein employs a traditional optical trap as well as an independent holographic optical trapping line, capable of creating and manipulating multiple traps. This allows for the creation of complex geometric arrangements of refractive particles while also permitting simultaneous high-speed, high-resolution measurements of the activity of biological enzymes.

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Biology

Flash-and-Freeze: A Novel Technique to Capture Membrane Dynamics with Electron Microscopy
Shuo Li *1,2, Sumana Raychaudhuri *1, Shigeki Watanabe 1,3
1Department of Cell Biology, Johns Hopkins School of Medicine, 2Department of Biochemistry and Molecular Biology, Johns Hopkins Bloomberg School of Public Health, 3Solomon H. Snyder Department of Neuroscience, Johns Hopkins School of Medicine

We developed a novel technique in electron microscopy, "flash-and-freeze," that enables the visualization of membrane dynamics with ms temporal resolution. This technique combines the optogenetic stimulation of neurons with high-pressure freezing. Here, we demonstrate the procedures and describe the protocols in detail.

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