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Analysis of Proteins by Immunoprecipitation - http://dps.ufl.edu/hansen/protocols/imp98.prt.htm

Category: Protein Protocols: Immunoprecipitation Protocols

Antibody-antigen complexes are removed from solution by addition of an insoluble form of an antibody binding protein such as Protein A, Protein G or second antibody. Immunoprecipitation protocols / methodology and technical background information. P.J. Ha - [Read Analysis of Proteins by Immunoprecipitation]

Immunoprecipitation Troubleshooting Guide PDF - http://www.stressgenbioreagents.com/literature/Troubleshooting-IP.pdf

Category: Protein Protocols: Immunoprecipitation Protocols: Immunoprecipitation Troubleshooting

Immunoprecipitation Troubleshooting Guide. Stressgen. Problem: Non-specific background, Specific Background, Specific antigen not detected. With Solutions to common IP problems. - [Read Immunoprecipitation Troubleshooting Guide PDF]

General Immunofluorescence Protocol - http://www.bcm.edu/rosenlab/protocols/generalIF.pdf

Category: Molecular Imaging: Immunofluorescence Microscopy

General Immunofluorescence Protocol. Rosen Lab. Deparaffinize and rehydrate sections, Antigen retrieval methods, Proteinase K Antigen Retrieval, Urea Antigen Retrieval, Blocking buffers, - [Read General Immunofluorescence Protocol]

Transient Transfection Into 293T Cells Protocol - http://www.cbrinstitute.org/labs/springer/protocols/jun_293T.html

Category: Transfection Protocols: Transient Transfection Protocols

Transient transfection into 293T cells is a convenient way to overexpress and obtain both cellular and extracellular (secreted or membrane) proteins. 293 is a human renal epithelial cell line which is transformed by adenovirus E1A gene product. 293T is a derivative which also express SV40 large T antigen, allowing episomal replication of plasmids containing the SV40 origin and early promoter region. They (both) have the unusual property of being highly transfectable. - [Read Transient Transfection Into 293T Cells Protocol]

Polyclonal Antibody Production Protocol - http://research.cm.utexas.edu/bkitto/Kittolabpage/Protocols/Immunology/PAb.html

Category: Antibody Protocols: Antibody Production Protocols

Protocol for polyclonal antibody production. Very useful for rapid and simple generation of antibodies for western blots, ELISA assays, and immunoprecipitation. Includes: Rabbit Immunization; Initial Preparation; Pre-bleed; Antigen Injection; Monitoring of Titer; Purification of Antibodies. - [Read Polyclonal Antibody Production Protocol]

Antigen Retrieval for Immunohistochemistry with Paraffin-Embedded Tissues Protocol - http://www.animal.ufl.edu/hansen/Protocols/Immunohistochemistry_paraffin.htm

Category: Immunohistochemistry Protocols: Antigen Retrieval Protocols

Protocol used to for immunohistochemistry on paraffin-embedded sections. Based on use of microwave energy to effect antigen retrieval. The immunohistochemistry procedure, is for use of Biomeda's HistoScan kit based on a streptavidin-peroxidase/biotinylated second antibody detection system with 3-amino, 9-ethylcarbazole (AEC) as chromogen. Undoubtedly, other kits or home-made reagents will also work . - [Read Antigen Retrieval for Immunohistochemistry with Paraffin-Embedded Tissues Protocol]

Fixing Attached Cells in Organic Solvents Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/21/pdb.prot4293

Category: Cell Culture Protocols: Cell Fixing Protocols

Organic solvents such as alcohols and acetone remove lipids and dehydrate cells, precipitating the proteins on the cellular architecture. Be aware that different antigens may be affected differently by the various solvents. If no previous data are available for your antigen, start with the 50/50 mixture. For tissue culture dishes, concentrations of acetone higher than 50% will destroy the integrity of the plastic. - [Read Fixing Attached Cells in Organic Solvents Protocol]

ANTIBODY PURIFICATION by affinity chromatography - http://mullinslab.ucsf.edu/protocols/html/ANTIBODY_PURIFICATION_by_affini.htm

Category: Chromatography Protocols: Immunoaffinity Chromatography Protocols

ANTIBODY PURIFICATION by affinity chromatography. By Beth, Mullins Lab UCSF. To affinity purify antibodies, generate lots of E. coli lysate that contains your antigen. If the protein can stand freeze thawing, then go ahead and purify the protein from e. coli lysate and keep it frozen until you need to couple it to a CH-sepharose column. - [Read ANTIBODY PURIFICATION by affinity chromatography]

Antibody Array Production and Analysis - http://www.stanford.edu/group/robinsonlab/protocols/protocols_tools.htm

Category: Antibody Microarray Protocols

Antibody Array Production and Data Analysis. Probing Antigen Arrays and washing/ Blocking. Great Protocols and information. Brian Kidd. Stanford. - [Read Antibody Array Production and Analysis]

Blocking Solutions Protocols - http://icg.cpmc.columbia.edu/cattoretti/Protocol/main_page_reagents.html#Blocking%20solutions

Category: Immunohistochemistry Protocols: Blocking Protocols

Protocols for blocking solutions. Includes: Endogenous peroxidase blocking solution (H2O2-Azide); Non-specific antibody background blocking solution: swine serum; Non-specific antibody background blocking solution: MILK; Enzymatic antigen retrieval solution; Unconjugated, biotin- and fluorochrome-conjugated antibody (primary) solutions; Unconjugated, biotin- and fluorochrome-conjugated antibody (secondary) solutions; Enzyme-conjugated secondary antibody solutions.; etc.. - [Read Blocking Solutions Protocols]

Antigen Retrieval PDF - http://www.dakousa.com/ishbantigenretr.pdf

Category: Immunology: Antigen Retrieval

Antigen Retrieval. Principles and Technique. A handbook of Immunochemical Staining Methods and techniques. Marc Key - [Read Antigen Retrieval PDF]

Isolation of Dendritic Cells Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E66369ADF1DF1962F01D3DA073183AB&objectid=6674A1FCBE1A61EE40A5E3EE8FB368F5

Category: Immunology: Mononuclear Cell Isolation Protocols

Presents two methods for preparing dendritic cells (DCs), a highly specialized type of antigen-presenting cell (APC). The first method involves the isolation of DCs from mouse spleen, resulting in a cell population that is highly enriched in accessory cell and APC function. A support protocol for collagenase digestion of splenocyte suspensions is described to increase the yield of dendritic cells. The second method involves generating large numbers of DCs from mouse bone marrow progenitor cells. - [Read Isolation of Dendritic Cells Protocol]

Antigen Retrieval - Immunohistochemistry - http://axon.med.harvard.edu/~cepko/protocol/mike/A2.html

Category: Immunology: Antigen Retrieval

Denaturing Solution and Procedure for AR. Mike Dyer. - [Read Antigen Retrieval - Immunohistochemistry]

Unmasking Hidden Epitopes Using the Microwave Oven Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/23/pdb.prot4529

Category: Immunohistochemistry Protocols: Antigen Retrieval Protocols

Several methods have been developed to "retrieve" antigens that have been masked by fixation. The principle behind using the microwave oven method described here is to use extended periods of heat to break some of the subcellular structures that block antibody access. Be aware that any of the antigen retrieval methods should be avoided wherever possible, because they may introduce artifactual false-positive staining. - [Read Unmasking Hidden Epitopes Using the Microwave Oven Protocol]

Antigen Retrieval for immunohistochemistry - http://herpesvirus.tripod.com/research/protoc.htm#Antigen%20Retrieval%20for

Category: Immunology: Antigen Retrieval

Antigen Retrieval for immunohistochemistry. Microwave method, Autoclave method. slides and staining. Janos Luka. Eastern Virginia. - [Read Antigen Retrieval for immunohistochemistry]

Double immunofluorescence staining for BCL-6 - http://icg.cpmc.columbia.edu/cattoretti/Protocol/Immunohistochemistry/DoubleIFstain.html

Category: Molecular Imaging: Immunofluorescence Microscopy

Double immunofluorescence staining for BCL-6. unfixed or acetone-fixed specimens. dewaxed, antigen retrieved slides - [Read Double immunofluorescence staining for BCL-6]

Antigen Retrieval Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/A2.html

Category: Immunohistochemistry Protocols: Antigen Retrieval Protocols

Protocol for antigen retrieval. - [Read Antigen Retrieval Protocol]

Immunoprecipitation: Preclearing the Lysate Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/23/pdb.prot4535

Category: Immunology: Immunoprecipitation Protocols: Protein Immunoprecipitation Protocols

To reduce backgrounds and to improve the signal-to-noise ratio, an antibody that does not recognize the antigen being studied can be added to the lysate and processed as for a normal immunoprecipitation. Any nonspecific proteins that might contaminate the final immunoprecipitation step are presumably removed with this irrelevant antibody. - [Read Immunoprecipitation: Preclearing the Lysate Protocol]

CD90 (Thy-1) Antibody Staining Protocol for Immunohistochemistry - http://www.ihcworld.com/_protocols/antibody_protocols/cd90_pharmingen.htm

Category: Immunohistochemistry Protocols: Antibody Staining Protocols

Protocol for CD90 (Thy-1) antibody staining for Immunohistochemistry. Includes: Primary Antibody; Antigen Retrieval; Detection Methods; Chromogen Substrate; Counterstain. - [Read CD90 (Thy-1) Antibody Staining Protocol for Immunohistochemistry]

General Immunohistochemistry Protocol - http://www.bcm.edu/rosenlab/protocols/generalIHC.pdf

Category: Immunohistochemistry Protocols

Protocol for general immunohistochemistry. Includes: Antigen retrieval methods; Sodium Citrate Antigen Retrieval; Proteinase K Antigen Retrieval; Urea Antigen Retrieval. - [Read General Immunohistochemistry Protocol]

Epitope Mapping by Competition Assay Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4277

Category: Antibody Protocols: Epitope Mapping Antibody Protocols

The simplest way to determine whether two monoclonal antibodies bind to distinct sites on a protein antigen is to carry out a competition assay. The assay can be used with antibodies that bind both conformational and linear epitopes, and it is most useful in the analysis of monoclonal antibody specificity because polyclonal sera typically recognize multiple different epitopes. - [Read Epitope Mapping by Competition Assay Protocol]

HIV-1 p24 Antigen Capture ELISA Protocol - http://ubik.microbiol.washington.edu/protocols/bl3/p24elisa.htm

Category: ELISA Protocols

Protocol for HIV-1 p24 antigen capture ELISA. - [Read HIV-1 p24 Antigen Capture ELISA Protocol]

Antigen Design and Sera Purification - http://www.sigmaaldrich.com/Brands/Sigma_Genosys/Custom_Antisera/Key_Resources/Antigen_Design.html

Category: Immunology: Antigen

Antigen Design and Sera Purification. Custom antisera. Sigma Aldrich. Peptide Selection and Design, Coupling Strategy # Selecting the Protein Carrier, Multiple Antigenic Peptides (MAPs), Choice of Host, Adjuvant, Immunization, & Sera Collection, Antisera Purification, Ammonium Sulfate Precipitation, Protein A/G, Immunoaffinity Purification. - [Read Antigen Design and Sera Purification]

Electron Microscopy of the Cytoskeleton of Cultured Cells - http://www.borisylab.northwestern.edu/pages/protocols/electmicrosctext.html

Category: Molecular Imaging: Electron Microscopy Protocols

The protocol for immunolabeling for electron microscopy depends mostly on the primary antibody and its ability to recognize antigen under particular ...Electron Microscopy of the Cytoskeleton of Cultured Cells. Boris Lab. - [Read Electron Microscopy of the Cytoskeleton of Cultured Cells]

AR Protease Digestion Method - http://www.chemicon.com/techsupp/Protocol/Protease.asp

Category: Immunology: Antigen Retrieval

Protease Antigen Retrieval (Trypsin) Protocol. Chemicon. - [Read AR Protease Digestion Method]

Articles (1-3 of 3)

In Vitro Translated Xenopus Mos Kinase Assay Protocol

In Vitro Translated Xenopus Mos Kinase Assay Protocol. In response to progesterone, immature Xenopus oocytes mature to eggs that can be fertilized. The Mos protein kinase is essential for oocyte maturation, most likely due to its ability to activate the MAP kinase cascade. This MAP kinase cascade eventually leads to the activation of Cdc2/cyclin B and entry into M phase. In this protocol, tagged Mos kinase is translated in vitro, immunopurified, and used in a kinase assay.

Antibodies Phage Expression Protocol Using 96-well Microtiter Plates

A Phage display expression protocol for antibody expression in 96-well microtiter plates.

Immobilized Antigen Phage Antibody Selection Protocol

A protocol for the selection of Phage Antibodies using Immobilized Antigen. This method describes the selection of antibodies from bacteriophage antibody libraries that recognize a specific antigen. The phage display library of antibody-displaying phage particles is exposed to antigen attached to a solid substrate (Nunc Immuno™ tubes). The phage particles with affinity for antigen bind to the immobilized antigen and are selected from the library of phage expressing antibodies.

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