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[[File:PnW.png|thumb|300x300px|Visit our Lab's Webpage at [[www.phagocytes.ca]]]]
[[File:PnW.png|thumb|300x300px|Visit our Lab's Webpage at [http://www.phagocytes.ca www.phagocytes.ca]]]
Welcome to the protocol wiki for the lab of [http://phagocytes.ca Dr. Bryan Heit], at the [http://www.uwo.ca University of Western Ontario]. This site contains the protocols, and other information, we frequently use in our lab.  Only laboratory members may edit pages, but anyone is welcome to use these protocols.  If you use these protocol, please cite us.  A link for generating citations can be found on the left side of the page.
Welcome to the protocol wiki for the lab of [http://phagocytes.ca Dr. Bryan Heit], at the [http://www.uwo.ca University of Western Ontario]. This site contains the protocols, and other information, we frequently use in our lab.  Only laboratory members may edit pages, but anyone is welcome to use these protocols.  If you use these protocol, please cite us.  A link for generating citations can be found on the left side of the page.


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= Index of Protocols =
= Index of Protocols =
 
{|width="100%"
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==== Lab Operations ====
===== Lab Operations =====
*[[Entrance Protocol]]
 
*[[Exit Protocol]]
*[[Entrance Protocol|Entrance Protocol]]  
*[[Labbook Guidelines|Guidelines for Proper Use of Laboratory Notebooks]]
*[[Exit Protocol|Exit Protocol]]  
*[[Saving Experimental Files|Proper Saving of Experimental Files]]
*[[Labbook Guidelines|Guidelines for Proper Use of Laboratory Notebooks]]  
*[[Saving Experimental Files|Proper Saving of Experimental Files]]  
*[[Setting Up Network Drives|Setting up Access to Network Drives & Wiki]]
*[[Setting Up Network Drives|Setting up Access to Network Drives & Wiki]]
<br>
===== General Protocols =====
*[[Common buffers|Common Buffers]]
*[[Cell Culture Guidelines|Cell Culture Guidelines]]
*[[Bacterial Growth Media|Bacterial Growth Media]]
*[[Primary Macrophage Culture|Primary Macrophage Culture]]
*[[Freezing and Thawing Cells|Freezing and Thawing Cells]]
*[[Raw Cell Transfection|Raw Cell Transfection]]
*[[J774 Cell Transfection|J774 Cell Transfection]]
*[[Antibiotics|Antibiotics]]
*[[Antibiotic Plates|Antibiotic Plates]]
*[[Agarose Gels|Agarose Gels]]
*[[Water Bath Antibiotic Solution|Water Bath Antibiotic Solution]]
<br>
===== Phagocytosis Protocols =====
*[[Opsonization|Opsonization]]
*[[Preparation of Silica-Magnetic Beads|Preparation of Silica-Magnetic Beads]]
*[[Synchronised Phagocytosis|Synchronised Phagocytosis]]
*[[Phagosome Isolation|Phagosome Isolation]]
*[[Primary Human Macrophages|Primary Human Macrophages]]
*[[Labelled E coli|Labelled E coli]]
*[[Inside-out Labelling of Bacteria|Inside-out Labelling of Bacteria]]
*[[Gentamicin Protection Assay|Gentamicin Protection Assay]]
<br> <br>
|
|
==== DNA/Cloning ====


===== DNA/Cloning =====
*[[Colony PCR]]
 
*[[PCR]]
*[[Colony PCR|Colony PCR]]<br>
*[[Digests]]
*[[PCR|PCR]]  
*[[Ligation]]
*[[Digests|Digests]]  
*[[Gibson Assembly]]
*[[Ligation|Ligation]]  
*[[Gibson Assembly|Gibson Assembly]]
*[[Quick 'n' Easy Competent E. coli|Quick 'n' Easy Competent ''E. coli'' (Dh5a)]]
*[[Quick 'n' Easy Competent E. coli|Quick 'n' Easy Competent ''E. coli'' (Dh5a)]]
*[[Competent e coli|Generating Competent ''E. coli'' (TFB, BL21 & ZYCY10P3S2T)]]
*[[Competent e coli|Generating Competent ''E. coli'' (TFB, BL21 & ZYCY10P3S2T)]]
*[[E. coli Transduction|Transducing ''E. coli'']]
*[[E. coli Transduction|Transducing ''E. coli'']]
*[[Transformation|Transformation]]
*[[Transformation]]
*[[Neon® Transfection System|Neon® Transfection System]]  
*[[Neon® Transfection System]]
*[[Generating Minicircles|Generating Minicircles]]  
*[[Generating Minicircles]]
<br>
|
==== Microscopy ====
*[[Competition of Charged Molecules with Lipophilic Cations]]
*[[Immunostaining|Fluorescent Immunostaining]]
*[[Inhibition of Focal Contact Signaling]]
*[[Single Particle Tracking]]
*[[Staining for GSD]]
*[[Reducing Photobleaching]]
*[[Acid Washing Coverslips]]
*[[Live Cell FRET]]
*[[3D Printed PDMS Chambers]]
|
|- style="vertical-align:top;"
|
==== General Protocols ====
*[[Common buffers|Common Buffers]]
*[[Cell Culture Guidelines]]
*[[Bacterial Growth Media]]
*[[Primary Macrophage Culture]]
*[[Freezing and Thawing Cells]]
*[[Raw Cell Transfection]]
*[[J774 Cell Transfection]]
*[[Antibiotics]]
*[[Antibiotic Plates]]
*[[Agarose Gels]]
*[[Water Bath Antibiotic Solution]]
|
==== Protein work ====
*[[Western Blotting]]
*[[Immunoprecipitation]]
*[[Fab preparation]]
*[[Fab Purification by FPLC|Fab purification using FPLC]]
*[[Updated FPLC Size Exclusion Procedure]]
*[[Coomassie Staining]]
*[[Receptor Cross-linking and Activation|Receptor activation by Cross-Linking]]
*[[Stripping & Reprobing Blots|Stripping &amp; Reprobing Blots]]
|
==== Lipids ====
*[[Asymmetric liposomes]]
*[[Lipid Coated Bead Preparation]]
*[[Lipid Extraction from Cells]]
*[[Lipisome and Lipid-Coated Beads]]
|
|- style="vertical-align:top;"
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==== Phagocytosis Protocols ====


===== Protein work =====
*[[Opsonization]]
 
*[[Preparation of Silica-Magnetic Beads]]
*[[Western Blotting|Western Blotting]]  
*[[Synchronised Phagocytosis]]
*[[Immunoprecipitation|Immunoprecipitation]]  
*[[Phagosome Isolation]]
*[[Fab preparation|Fab preparation]]  
*[[Primary Human Macrophages]]
*[[Fab Purification by FPLC|Fab purification using FPLC]]  
*[[Labelled E coli]]
*[[Updated FPLC Size Exclusion Procedure|Updated FPLC Size Exclusion Procedure]]
*[[Inside-out Labelling of Bacteria]]
*[[Coomassie Staining|Coomassie Staining]]  
*[[Gentamicin Protection Assay]]
*[[Receptor Cross-linking and Activation|Receptor activation by Cross-Linking]]  
*[[Preparation of Digestion-Tracking Bacteria]]
*[[Stripping & Reprobing Blots|Stripping &amp; Reprobing Blots]]  
|
 
==== Cell Biology ====
<br>
*[[Ablation of Recycling Endosomes]]
 
*[[Cell-Type Specific Transfection Protocols]]
===== Cell Biology<br> =====
 
*[[Ablation of Recycling Endosomes|Ablation of Recycling Endosomes]]  
*[[Cell-Type Specific Transfection Protocols|Cell-Type Specific Transfection Protocols]]  
*[[G418 & Puromycin Kill Curves|G418 &amp; Puromycin Kill Curves]]
*[[G418 & Puromycin Kill Curves|G418 &amp; Puromycin Kill Curves]]
*[[Apoptosis Detection with AnnexinV and PI|Apoptosis Detection with AnnexinV and PI]]  
*[[Apoptosis Detection with AnnexinV and PI]]
 
|
 
===== Microscopy =====
 
*[[Competition of Charged Molecules with Lipophilic Cations|Competition of Charged Molecules with Lipophilic Cations]]
*[[Immunostaining|Fluorescent Immunostaining]]
*[[Inhibition of Focal Contact Signaling|Inhibition of Focal Contact Signaling]]
*[[Single Particle Tracking|Single Particle Tracking]]
*[[Staining for GSD|Staining for GSD]]
*[[Reducing Photobleaching|Reducing Photobleaching]]
*[[Acid Washing Coverslips|Acid Washing Coverslips]]
*[[Live Cell FRET|Live Cell FRET]]
*[[3D Printed PDMS Chambers|3D Printed PDMS Chambers]]
 
<br>
 
===== Lipids<br> =====
 
*[[Asymmetric liposomes|Asymmetric liposomes]]<br>
*[[Lipid Coated Bead Preparation|Lipid Coated Bead Preparation]]<br>
*[[Lipid Extraction from Cells|Lipid Extraction from Cells]]<br>
*[[Lipisome and Lipid-Coated Beads|Lipisome and Lipid-Coated Beads]]<br>
 
<br>
 
 
 
<br>


|}
|}


= Links to More Protocols =
= Links to More Protocols =
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===== General Protocol Sites =====
==== General Protocol Sites ====
*[http://www.benchfly.com/ BenchFly] - Free Video Protocols  
*[http://www.benchfly.com/ BenchFly] - Free Video Protocols  
*[http://www.protocol-online.org/ Protocols Online] - large database of biology protocols  
*[http://www.protocol-online.org/ Protocols Online] - large database of biology protocols  
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*[http://www.thelabrat.com/protocols/reagents.shtml Common Buffer Recipes] @ TheLabRat  
*[http://www.thelabrat.com/protocols/reagents.shtml Common Buffer Recipes] @ TheLabRat  
*[http://www.rsc.org/Publishing/Journals/lc/Chips_and_Tips/index.asp Chips &amp; Tips] - Microfluidics protocols  
*[http://www.rsc.org/Publishing/Journals/lc/Chips_and_Tips/index.asp Chips &amp; Tips] - Microfluidics protocols  
|
|
===== Free Science Ebooks =====
==== Free Science Ebooks ====
*[https://www.gitbook.com/book/petebankhead/imagej-intro/details Analyzing fluorescence microscopy images with ImageJ]
*[https://www.gitbook.com/book/petebankhead/imagej-intro/details Analyzing fluorescence microscopy images with ImageJ]
*[http://www.imaging-git.com/applications/bioimage-data-analysis-0 Bioimage Data Analysis] - Free registration required
*[http://www.imaging-git.com/applications/bioimage-data-analysis-0 Bioimage Data Analysis] - Free registration required
|}
|}


= Useful Links =
= Useful Links =
 
{|width="100%"
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==== Molecular Biology Databases ====
===== Molecular Biology Databases =====
*[http://www.ncbi.nlm.nih.gov/gene Pubmed Gene] - Find gene information
 
*[http://www.ncbi.nlm.nih.gov/refseq/rsg/ Pubmed RefSeq] - Find reference sequences
*[http://www.ncbi.nlm.nih.gov/gene Pubmed Gene] - Find gene information<br>
*[http://www.uniprot.org/ Uniprot] - Find protein sequence &amp; structure
*[http://www.ncbi.nlm.nih.gov/refseq/rsg/ Pubmed RefSeq] - Find reference sequences<br>
*[http://hapmap.ncbi.nlm.nih.gov/ HapMap] - Find human polymophisms
*[http://www.uniprot.org/ Uniprot] - Find protein sequence &amp; structure<br>
*[http://www.ncbi.nlm.nih.gov/omim OMIM] - Find human gene-assocations
*[http://hapmap.ncbi.nlm.nih.gov/ HapMap] - Find human polymophisms<br>
*[http://www.pantherdb.org/ PANTHER] - Automated Protein Function Classification
*[http://www.ncbi.nlm.nih.gov/omim OMIM] - Find human gene-assocations<br>
*[http://useast.ensembl.org/index.html BioGrid] - Protein Interactions
*[http://www.pantherdb.org/ PANTHER] - Automated Protein Function Classification  
*[http://useast.ensembl.org/index.html Ensemble Genome] - Browse multiple genomes
*[http://useast.ensembl.org/index.html BioGrid] - Protein Interactions<br>
*[http://www.proteinatlas.org/ Human Protein Atlas] - Info onn gene exrpession, antibody's, etc
*[http://useast.ensembl.org/index.html Ensemble Genome] - Browse multiple genomes<br>
*[http://www.genecards.org/index.shtml Gene Cards] - Condenced information on genes
*[http://www.proteinatlas.org/ Human Protein Atlas] - Info onn gene exrpession, antibody's, etc<br>
*[http://www.ihop-net.org/UniPub/iHOP/ iHOP] - Information Hyperlinked Over Proteins
*[http://www.genecards.org/index.shtml Gene Cards] - Condenced information on genes<br>
*[http://www.hprd.org/index_html Human Protein Reference Database]
*[http://www.ihop-net.org/UniPub/iHOP/ iHOP] - Information Hyperlinked Over Proteins<br>
*[http://www.wwpdb.org/ PDB] - Protein Structures
*[http://www.hprd.org/index_html Human Protein Reference Database]<br>
*[http://genetics.bwh.harvard.edu/pph2/ PolyPhen2] - SNP Phenotype Predictor
*[http://www.wwpdb.org/ PDB] - Protein Structures<br>
*[http://sift.jcvi.org/ SIFT] - SNP Phenotype Predictor/db
*[http://genetics.bwh.harvard.edu/pph2/ PolyPhen2] - SNP Phenotype Predictor  
*[http://www.timetree.org/index.php TimeTree] - evolutionary divergence database
*[http://sift.jcvi.org/ SIFT] - SNP Phenotype Predictor/db  
|
*[http://www.timetree.org/index.php TimeTree] - evolutionary divergence database<br>  
==== Molecular Biology Tools ====
 
*[http://blast.ncbi.nlm.nih.gov/Blast.cgi NCBI Blast]
<br>
*[http://ca.expasy.org/ ExPASy Tools]
 
*[http://www.basic.northwestern.edu/biotools/oligocalc.html OligoCalc] - PCR primer Tm calculator
===== Journal Resources<br> =====
*[http://tools.neb.com/NEBcutter2/index.php NEB Cutter]
 
*[http://www.neb.com/nebecomm/tech_reference/restriction_enzymes/isoschizomers.asp NEB Isoschizomers]
*[http://www.ncbi.nlm.nih.gov/pubmed/ Pubmed]  
*[http://searchlauncher.bcm.tmc.edu/seq-util/Options/revcomp.html Reverse-Complement DNA Sequence]
*[http://www.ncbi.nlm.nih.gov/pmc/ Pubmed Central (USA)]  
*[http://www.insilico.uni-duesseldorf.de/Lig_Input.html Ligation Calculator]
*[http://pubmedcentralcanada.ca/ Pubmed Central (Canada)]  
*[http://www.addgene.org/ AddGene] - clone by e-mail!
*[http://scholar.google.com Google Scholar]  
*[http://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=npsa_pattinprot.html PattenProt] - search genomes for protein patterns
*[http://eigenfactor.org/ Eigenfactor] - free journal imapct scores  
*[http://workbench.sdsc.edu/ Biology Workbench]
 
*[http://www.ebi.ac.uk/Tools/sequence.html EMBL Sequence Tools]
<br>
*[http://www.ncbi.nlm.nih.gov/projects/gorf/ NCBI ORF Finder]
 
*[http://searchlauncher.bcm.tmc.edu/ BCM Search Launcher]
===== Genomics Resources<br> =====
*[http://swissmodel.expasy.org/ SWISS Model protein modeling]
*[http://searchlauncher.bcm.tmc.edu/seq-search/struc-predict.html Secondary Structure Prediction]
*[http://www.predictprotein.org/ PredictProtein] - Protein structure prediction
*[http://3d-alignment.eu/ STRAP] - Protein aligments with structure
|
===== Microscopy Tools =====
*[http://fbs.robarts.ca/ London Regional Microscopy Facility Bookings]
*[http://www.microscopyu.com/ Microscopy U] - Everything you want to know about microscopes
*[http://jcb.rupress.org/content/166/1/11.full Paper on Image Processing Standards] - How not to loose your job
*[http://www.invitrogen.com/site/us/en/home/support/Research-Tools/Fluorescence-SpectraViewer.html Fluorophore Spectra Viewer] at Life Technology
*[http://www.mcb.arizona.edu/ipc/fret/ Fluorescent Spectra Database] - FRET and other
*[http://www.mcb.arizona.edu/IPC/spectra_page.htm Yet More Spectra] - from Arizona University
*[http://www.confocal-microscopy.org/ www.confocal-microscopy.org] - Data on LSM methods and equipment
*[http://fiji.sc/wiki/index.php/Fiji FIJI] - <u>FREE</u> imageJ based image processing program
*[http://www.dspguide.com/pdfbook.htm Free] image processing textbook
*[http://www.archive.org/details/Lectures_on_Image_Processing Lectures on image processing]
*[http://vaa3d.org/ VAA3D] FREE viewer for large 3/4/5D datasets
|
|- style="vertical-align:top;"
|
==== Journal Resources ====
*[http://www.ncbi.nlm.nih.gov/pubmed/ Pubmed]
*[http://www.ncbi.nlm.nih.gov/pmc/ Pubmed Central (USA)]
*[http://pubmedcentralcanada.ca/ Pubmed Central (Canada)]
*[http://scholar.google.com Google Scholar]
*[http://eigenfactor.org/ Eigenfactor] - free journal imapct scores
|
==== ''In Vivo'' Tools ====
*[http://www.emouseatlas.org/emap/home.html EMAP] - Virtual mouse anatomy
*[http://phenome.jax.org/ Mouse phenome database] - mouse phenotypes<br>
|
==== Protease Tools ====
*[http://merops.sanger.ac.uk/ MEROPS] - Peptidase database
*[http://www.proteolysis.org/proteases PMAP] - Proteolysis Map
*[http://casbase.org/casvm/index.html CASVM] - Caspace substrate prediction
*[http://bioinf.gen.tcd.ie/casbah/ CASBAH] - Caspase cleaveage site database
|
|- style="vertical-align:top;"
|
==== Genomics Resources ====
*[http://www.gwascentral.org GWAS Central]
*[http://www.gwascentral.org GWAS Central]
|
==== Chemical Tools ====


|
*[http://www.chemspider.com/ ChemSpider] - General chemistry database
===== Molecular Biology Tools =====
*[http://pubchem.ncbi.nlm.nih.gov/ PubChem] - Chemical structure database
 
|
*[http://blast.ncbi.nlm.nih.gov/Blast.cgi NCBI Blast]
==== Lipid Tools ====
*[http://ca.expasy.org/ ExPASy Tools]<br>
*[http://www.lipidmaps.org/ Lipid Maps] - Lipidomics gateway at ''Nature''
*[http://www.basic.northwestern.edu/biotools/oligocalc.html OligoCalc] - PCR primer Tm calculator<br>
*[http://www.avantilipids.com/ Avanti Lipids] - Buy lipids
*[http://tools.neb.com/NEBcutter2/index.php NEB Cutter]<br>
*[http://www.neb.com/nebecomm/tech_reference/restriction_enzymes/isoschizomers.asp NEB Isoschizomers]<br>
*[http://searchlauncher.bcm.tmc.edu/seq-util/Options/revcomp.html Reverse-Complement DNA Sequence]<br>
*[http://www.insilico.uni-duesseldorf.de/Lig_Input.html Ligation Calculator]
*[http://www.addgene.org/ AddGene] - clone by e-mail!
*[http://npsa-pbil.ibcp.fr/cgi-bin/npsa_automat.pl?page=npsa_pattinprot.html PattenProt] - search genomes for protein patterns
*[http://workbench.sdsc.edu/ Biology Workbench]<br>
*[http://www.ebi.ac.uk/Tools/sequence.html EMBL Sequence Tools]<br>
*[http://www.ncbi.nlm.nih.gov/projects/gorf/ NCBI ORF Finder]<br>
*[http://searchlauncher.bcm.tmc.edu/ BCM Search Launcher]<br>
*[http://swissmodel.expasy.org/ SWISS Model protein modeling]<br>
*[http://searchlauncher.bcm.tmc.edu/seq-search/struc-predict.html Secondary Structure Prediction]<br>
*[http://www.predictprotein.org/ PredictProtein] - Protein structure prediction<br>
*[http://3d-alignment.eu/ STRAP] - Protein aligments with structure<br>
 
<br>
 
===== ''In Vivo'' Tools<br> =====
 
*[http://www.emouseatlas.org/emap/home.html EMAP] - Virtual mouse anatomy<br>
*[http://phenome.jax.org/ Mouse phenome database] - mouse phenotypes<br>
 
|  
===== Microscopy Tools =====
 
*[http://www.microscopyu.com/ Microscopy U] - Everything you want to know about microscopes<br>
*[http://jcb.rupress.org/content/166/1/11.full Paper on Image Processing Standards] - How not to loose your job
*[http://www.invitrogen.com/site/us/en/home/support/Research-Tools/Fluorescence-SpectraViewer.html Fluorophore Spectra Viewer] at Life Technology<br>
*[http://www.mcb.arizona.edu/ipc/fret/ Fluorescent Spectra Database] - FRET and other
*[http://www.mcb.arizona.edu/IPC/spectra_page.htm Yet More Spectra] - from Arizona University
*[http://www.confocal-microscopy.org/ www.confocal-microscopy.org] - Data on LSM methods and equipment<br>
*[http://fiji.sc/wiki/index.php/Fiji FIJI] - <u>FREE</u> imageJ based image processing program
*[http://www.dspguide.com/pdfbook.htm Free] image processing textbook
*[http://www.archive.org/details/Lectures_on_Image_Processing Lectures on image processing]<br>
*[http://vaa3d.org/ VAA3D] FREE viewer for large 3/4/5D datasets
 
<br>
 
===== Protease Tools =====
 
*[http://merops.sanger.ac.uk/ MEROPS] - Peptidase database
*[http://www.proteolysis.org/proteases PMAP] - Proteolysis Map
*[http://casbase.org/casvm/index.html CASVM] - Caspace substrate prediction
*[http://bioinf.gen.tcd.ie/casbah/ CASBAH] - Caspase cleaveage site database
 
<br>
 
===== Lipid Tools =====
 
*[http://www.lipidmaps.org/ Lipid Maps] - Lipidomics gateway at ''Nature''  
*[http://www.avantilipids.com/ Avanti Lipids] - Buy lipids  
 
<br>
 
===== Chemical Tools =====
 
*[http://www.chemspider.com/ ChemSpider] - General chemistry database
*[http://pubchem.ncbi.nlm.nih.gov/ PubChem] - Chemical structure database
 
<br>
 
<br>
 
|}
|}
<br>

Revision as of 12:52, 18 February 2021

Visit our Lab's Webpage at www.phagocytes.ca

Welcome to the protocol wiki for the lab of Dr. Bryan Heit, at the University of Western Ontario. This site contains the protocols, and other information, we frequently use in our lab.  Only laboratory members may edit pages, but anyone is welcome to use these protocols.  If you use these protocol, please cite us.  A link for generating citations can be found on the left side of the page.


Consult the User's Guide for information on using the wiki software.

Lab members:


Index of Protocols

Lab Operations

DNA/Cloning

Microscopy

General Protocols

Protein work

Lipids

Phagocytosis Protocols

Cell Biology

Links to More Protocols

General Protocol Sites

Free Science Ebooks

Useful Links

Molecular Biology Databases

Molecular Biology Tools
Microscopy Tools

Journal Resources

In Vivo Tools

Protease Tools

  • MEROPS - Peptidase database
  • PMAP - Proteolysis Map
  • CASVM - Caspace substrate prediction
  • CASBAH - Caspase cleaveage site database

Genomics Resources

Chemical Tools

Lipid Tools

Retrieved from "https://wiki.phagocytes.ca/index.php?title=Main_Page&oldid=85"