JoVE Bioengineering merges both physical and life sciences to understand and predict biological processes. Applying physical science tools to life science questions allow for the discovery of better technologies to measure, diagnose, and clinically treat disease.
1School of Dentistry, University of California, Los Angeles, 2School of Medicine, Clinical Nutrition, University of California, Los Angeles
Exosomes are microvesicular structures found within biofluids that potentially carry important disease discriminatory biomarkers. Here, a novel method is used to specifically extract exosomes and rapidly test the exosomal cargo for both RNA/protein targets following the disruption of exosomes using non-uniform electric cyclic square waves.
Published January 23, 2015. Keywords: Bioengineering, Exosome, Electrochemical sensors, Tumor biomarkers, Lung cancer, Salivary diagnostics
1Department of Nanomedicine, Houston Methodist Research Institute, 2MOE Key Laboratory of Bioinorganic and Synthetic Chemistry, School of Chemistry and Chemical Engineering, Sun Yat-sen University, 3Pediatrics Department of Union Hospital, Huazhong University of Science and Technology, 4CAS Key Laboratory for Biomedical Effects of Nanomaterials & Nanosafety, National Center for Nanoscience & Technology of China, 5Department of Medicine, Weill Cornell Medical College, 6Department of Cell and Developmental Biology, Weill Cornell Medical College
Delivery remains the main challenge for the therapeutic implementation of small interfering RNA (siRNA). This protocol involves the use of a multifunctional and biocompatible siRNA delivery platform, consisting of arginine and polyethylenimine grafted porous silicon microparticles.
Published January 15, 2015. Keywords: Bioengineering, Porous silicon, siRNA, Nanodelivery system, Cancer therapy, Transfection, Polycation functionalization
1Experimental Radiology, Institute of Diagnostic and Interventional Radiology I, Jena University Hospital, 2Department of Pharmaceutical Technology, Friedrich-Schiller-University Jena, 3Center for Electron Microscopy, Jena University Hospital
The use of fluorophores for in vivo imaging can be greatly limited by opsonization, rapid clearance, low detection sensitivity and cytotoxic effects on the host. Encapsulation of fluorophores in liposomes by film hydration and extrusion leads to fluorescence quenching and protection which enables in vivo imaging with high detection sensitivity.
Published January 5, 2015. Keywords: Bioengineering, Drug-delivery, Liposomes, Fluorochromes, Fluorescence-quenching, Optical imaging, Inflammation
1Angiogenesis Laboratory, Department of Ophthalmology, Harvard Medical School, Massachusetts Eye & Ear Infirmary, 2St. Elizabeth's Medical Center
Here, we present a protocol that allows the investigator to assess leukocyte recruitment dynamics ex vivo by connecting a chamber coated with endothelial-derived adhesion molecules to the circulatory system of a mouse. This method offers significant advantages since it allows for leukocyte assessment under relative biological conditions.
Published December 30, 2014. Keywords: Bioengineering, Leukocyte recruitment, endothelium, flow chamber, adhesion, velocity, rolling, integrins, ex vivo
1Department of Chemistry and Biochemistry, University of Notre Dame, 2Harper Cancer Research Institute, University of Notre Dame
Immortalized cancer cell lines can be grown as 3D cell cultures, a valuable model for biological research. This protocol describes mass spectrometry imaging of 3D cell cultures, including improvements in the sample preparation platform. The goal of this protocol is to instruct users to prepare 3D cell cultures for mass spectrometry imaging analysis.
Published December 5, 2014. Keywords: Bioengineering, 3D cell culture, mass spectrometry, imaging, cell culture, sample preparation, spheroids
1Department of Epidemiology, School of Public Health, The University of Michigan, 2Centre for Oral Health Research, School of Dental Sciences, Newcastle University
The goal of this methods paper is to describe the use of a microfluidic system for the development of multi-species biofilms that contain species typically identified in human supragingival dental plaque. Methods to describe biofilm architecture, biofilm viability, and an approach to harvest biofilm for culture-dependent or culture-independent analyses are highlighted.
Published December 1, 2014. Keywords: Bioengineering, Dental plaque, biofilm, confocal laser scanning microscopy, three-dimensional structure, pyrosequencing, image analysis, image reconstruction, saliva, modeling, COMSTAT, IMARIS, IMAGEJ, multi-species biofilm communities.
1BioNano Laboratory, School of Engineering, University of Guelph
Here, we present a protocol explaining the use of Kelvin probe force microscopy as a tool for generating high resolution nano-scale surface potential maps. This tool was applied to assess the role of surface potential on the binding capacity of microorganisms to substrate surfaces.
Published November 28, 2014. Keywords: Bioengineering, Kelvin probe force microscopy, atomic force microscopy, surface potential, stainless steel, microbial attachment, bacterial biofilms, methicillin-resistant Staphylococcus aureus
1Institute for Biological and Medical Imaging (IBMI), Helmholtz Zentrum München, 2Faculty of Medicine, Technische Universität München
We provide herein a detailed description of the experimental protocol for imaging with a newly developed hand-held optoacoustic (photoacoustic) system for three-dimensional functional and molecular imaging in real time. The demonstrated powerful performance and versatility may define new application areas of the optoacoustic technology in preclinical research and clinical practice.
Published November 4, 2014. Keywords: Physiology, Optoacoustic tomography, photoacoustic imaging, hand-held probe, volumetric imaging, real-time tomography, five dimensional imaging, clinical imaging, functional imaging, molecular imaging, preclinical research
1Department of Biomedical Engineering, University of Wisconsin, Madison, 2Department of Mechanical Engineering, University of Wisconsin, Madison
Cell culture substrates functionalized with microscale patterns of biological ligands have immense utility in the field of tissue engineering. Here, we demonstrate the versatile and automated manufacture of tissue culture substrates with multiple, micropatterned poly(ethylene glycol) brushes presenting orthogonal chemistries that enable spatially precise and site-specific immobilization of biological ligands.
Published October 31, 2014. Keywords: Bioengineering, Robotic microcontact printing, R-μCP, click chemistry, surface chemistry, tissue engineering, micropattern, advanced manufacturing
1Department of Otolaryngology, University of California, San Francisco, 2Department of Chemistry, University of California, Berkeley, 3Materials Science Division, Lawrence Berkeley National Laboratory, 4Department of Pharmaceutical Chemistry, University of California, San Francisco, 5Tetrad Graduate Program, University of California, San Francisco, 6Center for Systems and Synthetic Biology, University of California, San Francisco, 7Chemistry and Chemical Biology Graduate Program, University of California, San Francisco
We provide detailed instructions for the preparation of monovalent targeted quantum dots (mQDs) from phosphorothioate DNA of defined length. DNA wrapping occurs in high yield, and therefore, products do not require purification. We demonstrate the use of the SNAP tag to target mQDs to cell-surface receptors for live-cell imaging applications.
Published October 23, 2014. Keywords: Bioengineering, monovalent quantum dots, single particle tracking, SNAP tag, steric exclusion, phosphorothioate, DNA, nanoparticle bioconjugation, single molecule imaging