Hancock Lab Protocols

 

Dr R. E.W. (Bob) Hancock, OC, FRSC

Canada Research Chair and Professor

Department of Microbiology and Immunology, UBC

 

 

 

Antibiotics and Antimicrobial Peptides

• Beta-lactamase induction (crude preparation)

• Dinitrophenylation Assay For Polymixin B

• MIC Determination by Microtitre Broth Dilution Method

• Modified MIC Method for Cationic Antimicrobial Peptides

• Penicillin Binding Protein Assay

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Bacterial Genetics

• Alkaline Lysis Plasmid Miniprep

• Blotting and Hybridization with Sodium Phosphate

• Blunt End Cloning of PCR Products

• Chromosomal DNA Isolation

• Cosmid Library

• DNA Isolation from Agarose Gels with DEAE Paper

• Grid for 100 Clones

• Isolation of Chromosomal DNA

• Isolation of DNA from Agarose Gels with DEAE Paper

• Isolation pf Phage Resistant Mutants

• Lambda-TnphoA Mutagenesis of Cloned Genes in E. Coli

• M13 Single-Stranded Template Preparation

• Phage Resistant Mutant Isolation

• Preparation of M13 Single-Stranded Template

• Quick Isolation of DNA from Agarose Gels

• Slot Lysis

• Southern Blot

• Transposon Mutagenesis of Pseudomonas Aeruginosa

• Transformation of Helicobacter Pylori

• Biparental Mating

• CaCl₂ Transformation of E. coli

• Colony Blotting with ³²P

• Colony Immunoblot

• CsCl-Ethidium Bromide Gradient Plasmid Preparations

Protein Purification and Gel Electrophoresis

• Acetylation (or Succinylation) of Amino Groups on Proteins

• Acid-Urea Gel Electrophoresis

• Crosslinking of Intact Cells

• Crosslinking of Proteins

• Crosslinking of Whole Cells

• Electroblotting on to PVDF (Immobilon) Membrane

• Estimation of Amino Groups Using TNBS

• Isoelectric Focussing of Membrane Proteins by Slab Gel Method

• Isolation of Outer Membrane Protein from P. aeruginosa With Octyl-POE

• Isolation of Outer Membrane Protein With Zwittergent 3-14

• Lipid A Preparation

• Outer Membrane Protein Solubilization from P. aeruginosa with Octyl-POE

• Outer Membrane Protein Solubilization with Zwittergent 3-14

• Seperation of Outer and Inner Membranes

• Porin F Purification

• Protein and LPS Gels

• Protein Assay for Detergent Solubilized Proteins

• Whole Cell Protein and LPS Gels

Liposome Methods

• Liposome Preparation

• Liposome Swelling Assay

• Phospholipid Extraction

• Planar Lipid Bilayer Experiment

• Planer (Black) Lipid Bilayer Apparatus

Outer Membranes

• Cell Envelope Preparation / Solubilization

• Crosslinking of Intact Cells

• Crosslinking of Proteins

• Crosslinking of Whole Cells

• Dansyl-Polymyxin Synthesis

• Dinitrophenylation Assay For Polymixin B

• Inner Membrane Permeability Assay(ONPG Assay)

• KDO Assay

• Liposome Swelling Assay

• LPS Isolation (Darveau-Hancock Method)

• Methylation of Fatty Acids (Kropinski Method)

• Nitrocefin Assay on Whole Cells for Outer Membrane Permeability

• Nitrocefin Assay on Whole Cells for Outer Membrane Permeability Assessment

• NPN Uptake Assay / Flourescence Measurements

• Osmotic Shock Procedures

• Outer Membrane Preparation - One Step Sucrose Gradient Procedure

• Outer Membrane Protein Solubilization from P. aeruginosa with Octyl-POE

• Outer Membrane Protein Solubilization with Zwittergent 3-14

• Permeabilization of Whole Cells to Lysozyme

• Phase Partitioning of Cells

• Phosphate Uptake Assay

• Planar Lipid Bilayer Experiment

• Protease Digestion of Outer Membranes

• Planar (Black) Lipid Bilayer Apparatus

• Planar Lipid Bilayer Experiment

• Synthesis of Dansyl-Polymyxin

• French Press

Biochemical Assays

• Acetylation (or Succinylation) of Amino Groups on Proteins

• Alkaline Phosphatase C Assay for P. aeruginosa

• Alkaline Phosphatase and Phospholipase C Assays

• Carbohydrate Assays

• Dansyl-Polymyxin Synthesis

• Dehydrogenase Assays

• Dinitrophenylation Assay For Polymixin B

• Estimation of Amino Groups Using TNBS

• Glucosamine Rapid Assay

• KDO Assay

• Muramic Acid Assay

• Penicillin Binding Protein Assay

• Phenol Sulphuric Acid Carbohydrate Assay

• Phosphate Assay

• Phosphate Uptake Assay

• Whole Cell Phosphate Uptake Assay

Microbiological Techniques

• Freezing Bacterial Cells

• Indirect Immunofluorescence

• Methionine Labelling

• Oxidase Test

• Phage Propagation

• Stains

• TSBD Trypticase Soy Broth Dialysate

Host-Pathogen Interactions

• In Vivo Chamber Model

• Tissue Culture - Storage of Cell Lines

• Tissue Culture - Thawing Cells from Liquid Nitrogen

Functional Genomics

• Transposon Mutagenesis of Pseudomonas Aeruginosa

Immunology Based Methods

• Agglutination Assays

• Double Immunodiffusion

• Fusion Protocol

• Immunoprecipitation

• Tissue Culture - Storage of Cell Lines

• Tissue Culture - Thawing Cells from Liquid Nitrogen

• Endotoxin Assay

• Enzyme Linked Immunosorbent Assay