Intranet

Documents for New Lab Members

• Hsu Lab roadmap (expectations - UT) — read first
• Hsu Lab policy document
• Hsu Lab roadmap (expectations - UVA)
• Training plan — Stage 1 (Fall after joining)
• Training plan — Stage 2 (Spring after joining)
• IDP forms for performance reviews

LC-MS Training

• Introduction to Proteomics (Hsu Lab)
• Introduction to Data Analysis (Hsu Lab)
• Introduction to Proteomics (Broad Institute)

Operation / Maintenance on the QE+

• General overview from Thermo (PDF)
• Removing the ion transfer tube (video)
• Removing the S-lens (video)
• Cleaning the S-lens (video)
• Reinstalling the S-lens (video)

Mass Spec Standards (QC)

• Preparation of PRTC MS Standard
• Preparation of HeLa tryptic peptide digest

Tutorials

• Tissue Culture Basics
• Minimizing tissue culture contaminations
• Traditional Cloning Basics
• Bacterial Transformation and Competent Cells
• Bacterial Transformation Workflow
• Polymerase Chain Reaction (PCR)

Databases

• Lipid Maps (great website for intro to lipids)
• Hsu Lab Proteomics Database (IP2)
• Hsu Lab Synology (Data network attached server)
• Hsu Lab Synology (LC-MS data server)
• Mass Spectrometry Adduct Calculator

Safety Information

• UVa EHS
• Chemical Safety
• Chemical Waste (includes Sharps)

Primary Macrophage and Dendritic Cell Differentiation

• BMDM Macrophage differentiation protocol
• BMDC Dendritic cell differentiation protocol

Standard Operating Protocols

• How to keep a lab notebook (READ FIRST)
• Preparing SILAC media
• Agar Plates
• Bacterial Glycerol Stocks
• Sample Prep for LC-MS proteomics (click chemistry, on-bead trypsin digestion, target ID)
• Sample Prep for LC-MS proteomics (click chemistry, site of binding)
• Treatment of cells with compounds
• Preparing cells for cryostorage in N2 cell dewar
• Transforming E. coli (chemically competent)
• Gel-based ABPP (click chemistry protocol)
• Hsu Lab general LC-MS procedures (read before performing ANY mass spec experiments)
• Subculturing cells
• Lipid extraction of cells/tissues for LC-MS lipidomics
• Harvesting tissues from mice
• Processing cells/tissues for ABPP without sonication (optimal DAGL activity)
• Processing cells/tissues for ABPP with sonication (standard technique)
• Transient transfections
• ATP Acyl Phosphate Gel-Based ABPP Analysis
• ATP Acyl Phosphate LC-MS ABPP Analysis
• ATP Acyl Phosphate LC-MS ABPP Analysis (SILAC)
• Preparing Angiotensin/Vasoactive intestinal peptide standard stocks for LC-MS
• Filter Aided Sample Preparation (FASP) Method for Proteomics analysis
• ADP-glo substrate assay
• T cell activation (CD3/CD28) and anergy (CD3 only) assay
• TMT site of binding protocol
• General Buffers for lab

Molecular Biology / Cloning

• Preparing electrocompetent XL1 blue E. coli cells for transformation
• Site-directed mutagenesis
• E. coli electroporation and propagation of plasmid DNA
• Gateway system — transfer from donor to expression plasmid
• Sequencing to confirm successful cloning
• Serial Cloner download (for viewing lab plasmid maps)
• Excel file of plasmids and primers in lab
• Plasmid maps (Serial Cloner format)

Gel Electrophoresis

• Handcasting polyacrylamide long gels (protocol)
• Handcasting polyacrylamide long gels (reagent order list)

Software

• Pinpoint
• Tracefinder
• Skyline Proteomics Analysis Software

Capillary Column Pulling

• Sutter P2000 website
• Sutter P200 manual
• Packing Capillary column protocol
• Normal phase column parameters
• Reverse phase column parameters