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ShanghaiTech University

10 ARTICLES PUBLISHED IN JoVE

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Immunology and Infection

Scalable High Throughput Selection From Phage-displayed Synthetic Antibody Libraries
Shane Miersch 1,2, Zhijian Li 1,2, Rachel Hanna 1,2, Megan E. McLaughlin 1,2, Michael Hornsby 1,3, Tet Matsuguchi 1,3, Marcin Paduch 1,4, Annika Sääf 1,4, Jim Wells 1,3, Shohei Koide 1,4, Anthony Kossiakoff 1,4, Sachdev S. Sidhu 1,2
1The Recombinant Antibody Network, 2The Banting and Best Department of Medical Research, University of Toronto, 3Antibiome Center, University of California, San Francisco at Mission Bay, 4Department of Biochemistry and Molecular Biology, The University of Chicago

A method is described with visual accompaniment for conducting scalable, high throughput selections from phage-displayed combinatorial synthetic antibody libraries against hundreds of antigens simultaneously. Using this parallel approach, we have isolated antibody fragments that exhibit high affinity and specificity for diverse antigens that are functional in standard immunoassays.

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Biology

Direct Protein Delivery to Mammalian Cells Using Cell-permeable Cys2-His2 Zinc-finger Domains
Thomas Gaj *1, Jia Liu *1,2
1Departments of Chemistry and Cell and Molecular Biology, The Scripps Research Institute, 2Shanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University

Zinc-finger domains are intrinsically cell-permeable and capable of mediating protein delivery into a broad range of mammalian cell types. Here, a detailed step-by-step protocol for implementing zinc-finger technology for intracellular protein delivery is presented.

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Neuroscience

Optogenetic Functional MRI
Peter Lin 1, Zhongnan Fang 2, Jia Liu 1, Jin Hyung Lee 1,2
1Neurology and Neurological Sciences, Stanford University, 2Electrical Engineering, Neurology and Neurological Sciences, Stanford University

This protocol describes the steps and data analysis required to successfully perform optogenetic functional magnetic resonance imaging (ofMRI). ofMRI is a novel technique that combines high-field fMRI readout with optogenetic stimulation, allowing for cell type-specific mapping of functional neural circuits and their dynamics across the whole living brain.

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Developmental Biology

Whole-cell Patch-clamp Recordings of Isolated Primary Epithelial Cells from the Epididymis
Bao Li Zhang 1,2,3, Da Yuan Gao 1,2,3, Xiao Xu Zhang 1,3, Shuo Shi 4, Winnie Shum 1
1School of Life Science and Technology, ShanghaiTech University, 2Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 3University of Chinese Academy of Sciences, 4Shanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University

We present a protocol that combines cell isolation and whole-cell patch-clamp recording to measure the electrical properties of the primary dissociated epithelial cells from the rat cauda epididymides. This protocol allows for investigation of the functional properties of primary epididymal epithelial cells to further elucidate the physiological role of the epididymis.

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Bioengineering

Production of Chemicals by Klebsiella pneumoniae Using Bamboo Hydrolysate as Feedstock
Dong Wei 1,2, Jinjie Gu 1, Zhongxi Zhang 1, Chenhong Wang 1, Dexin Wang 1, Chul Ho Kim 3, Biao Jiang 1, Jiping Shi 1,4, Jian Hao 1
1Lab of Biorefinery, Shanghai Advanced Research Institute, Chinese Academy of Sciences, 2University of Chinese Academy of Sciences, 3Biorefinery Research Center, Jeonbuk Branch Institute, Korea Research Institute of Bioscience & Biotechnology (KRIBB), 4School of Life Science and Technology, ShanghaiTech University

Bamboo powder was pretreated with NaOH and enzymatically hydrolyzed. The hydrolysate of bamboo was used as the feedstock for 2,3-butanediol, R-acetoin, 2-ketogluconic acid, and xylonic acid production by Klebsiella pneumoniae.

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Immunology and Infection

Construction of Synthetic Phage Displayed Fab Library with Tailored Diversity
Ganggang Huang *1, Zhenwei Zhong *1, Shane Miersch 2,3, Sachdev S. Sidhu 1,2,3, Shin-chen Hou 1, Donghui Wu 1
1Laboratory of Antibody Engineering, Shanghai Institute for Advanced Immunochemical Studies, ShanghaiTech University, 2Banting and Best Department of Medical Research, Terrence Donnelly Center for Cellular and Biomolecular Research, University of Toronto, 3Department of Molecular Genetics, Terrence Donnelly Center for Cellular and Biomolecular Research, University of Toronto

This protocol describes a detailed procedure for the construction of a phage-displayed synthetic antibody library with tailored diversity. Synthetic antibodies have broad applications from basic research to disease diagnostics and therapeutics.

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Genetics

In Vitro Biochemical Assays using Biotin Labels to Study Protein-Nucleic Acid Interactions
Lina Yu *1, Wenxiu He *1, Jie Xie *1, Rui Guo *1, Juan Ni 2, Xia Zhang 3, Quishi Xu 1, Caifeng Wang 1, Qiuling Yue 1, Fangfang Li 1, Mengcheng Luo 4, Bo Sun 3, Lan Ye 1, Ke Zheng 1
1State Key Laboratory of Reproductive Medicine, Nanjing Medical University, 2The Affiliated Hospital of Hangzhou Normal University, 3School of Life Science and Technology, ShanghaiTech University, 4Department of Tissue and Embryology, School of Basic Medical Sciences, Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan University

Presented here are protocols for in vitro biochemical assays using biotin labels that may be widely applicable for studying protein-nucleic acid interactions.

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Chemistry

Quantitative SERS Detection of Uric Acid via Formation of Precise Plasmonic Nanojunctions within Aggregates of Gold Nanoparticles and Cucurbit[n]uril
Weng-I Katherine Chio 1, Gemma Davison 1, Tabitha Jones 1, Jia Liu 1, Ivan P. Parkin 1, Tung-Chun Lee 1,2
1Department of Chemistry, University College London (UCL), 2Institute for Materials Discovery, University College London (UCL)

A host-guest complex of cucurbit[7]uril and uric acid was formed in an aqueous solution before adding a small amount into Au NP solution for quantitative surface-enhanced Raman spectroscopy (SERS) sensing using a modular spectrometer.

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Biochemistry

Purification of Endogenous Drosophila Transient Receptor Potential Channels
Jia Liu *1, Yuyang Liu *1, Weidi Chen 1, Yuzhen Ding 1, Xiaoru Lan 1, Wei Liu 1
1Shenzhen Key Laboratory for Neuronal Structural Biology, Biomedical Research Institute, Shenzhen Peking University-The Hong Kong University of Science and Technology Medical Center

Based on the assembling mechanism of the INAD protein complex, in this protocol, a modified affinity purification plus competition strategy was developed to purify the endogenous Drosophila TRP channel.

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Cancer Research

An R-Based Landscape Validation of a Competing Risk Model
Haiping Lin *1,3, Hongjuan Zheng *2, Chenyang Ge 3, Lixian Ling 2, Ruihua Yin 2, Qinghua Wang 2, Xia Zhang 2, Shishi Zhou 2, Xiayun Jin 2, Xifeng Xu 2, Jianfei Fu 2
1Department of Hepatobiliary Surgery, Affiliated Jinhua Hospital, Zhejiang University School of Medicine, 2Department of Medical Oncology, Affiliated Jinhua Hospital, Zhejiang University School of Medicine, 3Department of Colorectal Surgery, Affiliated Jinhua Hospital, Zhejiang University School of Medicine

The present protocol describes codes in R for evaluating the discrimination and calibration abilities of a competing risk model, as well as codes for the internal and external validation of it.

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