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University of California, San Diego

6 ARTICLES PUBLISHED IN JoVE

Biology

Using Unfixed, Frozen Tissues to Study Natural Mucin Distribution

Miriam Cohen¹, Nissi M. Varki¹, Mark D. Jankowski², Pascal Gagneux¹

¹Department of Cellular and Molecular Medicine, University of California, San Diego , ²Biosecurity and Public Health, Los Alamos National Laboratory

Unfixed frozen tissue samples embedded in Optimal Cutting Temperature medium (OCT) can be used to study natural distribution and glycosylation of secreted mucus. In this approach tissue processing is minimal and the natural presentation of glycolipids, mucins and glycan-epitopes is preserved. Tissue sections can be analyzed by immunohistochemistry using fluorescence or chromogenic detection.

Medicine

Sampling Human Indigenous Saliva Peptidome Using a Lollipop-Like Ultrafiltration Probe: Simplify and Enhance Peptide Detection for Clinical Mass Spectrometry

Wenhong Zhu¹, Richard L. Gallo^2,3, Chun-Ming Huang^2,3,4

¹Sanford-Burnham Medical Research Institute, ²Division of Dermatology, University of California, San Diego , ³VA San Diego Healthcare Center, ⁴Moores Cancer Center, University of California, San Diego

Considering saliva sampling for future clinical application, a lollipop-like ultrafiltration (LLUF) probe was fabricated to fit in the human oral cavity. Direct analysis of undigested saliva by NanoLC-LTQ mass spectrometry demonstrated the ability of LLUF probes to remove large proteins and high abundance proteins, and make low-abundant peptides more detectable.

Bioengineering

Density Gradient Multilayered Polymerization (DGMP): A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Shivanjali Joshi-Barr¹, Jerome V. Karpiak², Yogesh Ner¹, Jessica H. Wen³, Adam J. Engler³, Adah Almutairi^1,2

¹Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego , ²Biomedical Sciences Program, University of California, San Diego , ³Department of Bioengineering, University of California, San Diego

Here we describe a unique strategy for creating biocompatible, layered matrices with continuous interfaces between distinct layers for tissue engineering. Such a scaffold could provide an ideal customizable environment to modulate cell behavior by various biological, chemical or mechanical cues

Biology

A High-throughput Method for Measurement of Glomerular Filtration Rate in Conscious Mice

Timo Rieg^1,2

¹Division of Nephrology-Hypertension, Department of Medicine, University of California, San Diego , ²San Diego VA Healthcare System

Measurement of glomerular filtration rate (GFR) is the gold standard for kidney function assessment. Here we describe a high-throughput method which allows the determination of GFR in conscious mice by using a single bolus injection, determination of fluorescein isothiocyanate (FITC)-inulin in plasma and calculation of GFR by a two-phase exponential decay model.

Immunology and Infection

Measuring Growth and Gene Expression Dynamics of Tumor-Targeted S. Typhimurium Bacteria

Tal Danino^*1, Arthur Prindle^*2, Jeff Hasty^2,3,4, Sangeeta Bhatia^1,5,6,7,8

¹Health Sciences and Technology, Massachusetts Institute of Technology, ²Department of Bioengineering, University of California, San Diego , ³Biocircuits Institute, University of California, San Diego , ⁴Molecular Biology Section, Division of Biological Science, University of California, San Diego , ⁵Broad Institute of Harvard and MIT, ⁶Department of Medicine, Brigham and Women's Hospital, ⁷Electrical Engineering and Computer Science and David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, ⁸Howard Hughes Medical Institute

The goal of these experiments is to generate quantitative time-course data on the growth and gene expression dynamics of attenuated S. typhimurium bacterial colonies growing inside tumors. This video covers tumor cell preparation and implantation, bacteria preparation and injection, whole-animal luminescence imaging, tumor excision, and bacterial colony counting.

Biology

Rapid, Affordable, and Uncomplicated Production of Bacterial Cell-free Lysate

Robert M. Cooper^*1, Taishi Tonooka^*1,2, Andriy Didovyk^*1,3, Jeff Hasty^1,4,5

¹Biocircuits Institute, University of California, San Diego, ²Present address, Kyoto Institute of Technology, ³Present address, Vertex Pharmaceuticals, ⁴Department of Bioengineering, University of California, San Diego, ⁵Molecular Biology Section, University of California, San Diego

This protocol describes a rapid and simple method to produce bacterial lysate for cell-free gene expression, using an engineered strain of Escherichia coli and requiring only standard laboratory equipment.