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Combined DNA In Situ Hybridization and Immunocytochemistry Protocol - http://www.roche-applied-science.com/PROD_INF/MANUALS/InSitu/pdf/ISH_127-134.pdf

Category: Cytogenetics Protocols: In Situ Hybridization Protocols

Protocol for combined DNA in situ hybridization and immunocytochemistry for the simultaneous detection of nucleic acid sequences, proteins, and incorporated BrdU in cell preparations. Includes: Cell preparations and BrdU labeling; Detection of antigen by immunocytochemistry (ICC); Visualization of ICC antigen; -Gal-BCIG reaction (for producing a blue precipitate visible under brightfield microscopy); Cell processing for in situ hybridization; In situ hybridization (ISH); etc... - [Read Combined DNA In Situ Hybridization and Immunocytochemistry Protocol]

Constructing and Expressing GFP Fusion Proteins - http://www.cshprotocols.org/cgi/content/abstract/2006/30/pdb.prot4649

Category: Reporter Gene Assays: GFP Assay Protocols

The following protocol can be used for the development of stable cell lines expressing GFP fusion proteins. Although optimal transfection procedures (e.g., calcium phosphate, electroporation, or FuGENE 6 [Roche Applied Science]) vary depending on cell type, this general transfection procedure has been successful for stable transfection of HeLa, A-431, U2OS, BHK, and HT1080 cells. - [Read Constructing and Expressing GFP Fusion Proteins]

Coupling of Proteins to IDC UltraClean Chloromethyl Latex Protocol - http://www.idclatex.com/body_bgrounder-superactive-protocol-11.asp

Category: Microsphere Analysis Protocols

Protocol for coupling of proteins to IDC UltraClean Chloromethyl latex. - [Read Coupling of Proteins to IDC UltraClean Chloromethyl Latex Protocol]

Cryosectioning Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/C7.html

Category: Immunohistochemistry Protocols: Cryosectioning Protocols

Protocol for cryosectioning. While the timing of the various steps in this protocol are probably not critical, process the tissue all at once to ensure that RNA and/or proteins do not get degraded. Includes: 20% Paraformaldehyde/4% Paraformaldehyde-PBS; Sucrose/PBS. - [Read Cryosectioning Protocol]

Culture of Endometrial Explants and Peri-implantation Conceptuses to Monitor Synthesis and Secretion - http://www.animal.ufl.edu/hansen/protocols/endocult.htm

Category: Cell Culture Protocols

Protocol used to study secretion of proteins and prostaglandins by endometrium from the cow, ewe, mare, bitch and other species. The technique is also useful for culture of peri-implantation conceptuses and placental tissues for metabolic labelling studies and to obtain conceptus secretory proteins for biological studies.The medium used is called Pig MEM, which is a modified minimum essential medium supplemented with non-essential amino acids, vitamins, insulin and additional glucose. - [Read Culture of Endometrial Explants and Peri-implantation Conceptuses to Monitor Synthesis and Secretion]

Culturing of Insect Cells and Production of Baculovirus Expressed Recombinant Proteins in 2.8 Liter - http://www.hyclone.com/pdf/procedure_insect_fernbach.pdf

Category: Cell Culture Protocols: Insect Cell Culture Protocols

The use of Ferbach flasks is an ideal method to amplify the cells and virus inoculum for the bioreactor. The method is also very practical for producing sufficient amounts of various proteins for feasibility studies using HyQ© SFX-Insect™ medium. The large surface area in this vessel enables sufficient aeration and, therefore, the cells, when they reach high density, are not oxygen depleted. - [Read Culturing of Insect Cells and Production of Baculovirus Expressed Recombinant Proteins in 2.8 Liter]

Cyanine Dye (Maleimide) Protein Labeling Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000150.pdf?pid=PP00000150

Category: Fluorescent Dyes Protocols

Cyanine dye reagents are useful as fluorescent labels for proteins. This protocol has been designed to label the thiol group on cysteine using Cy3 or Cy5 minimal maleimide labeling dyes. - [Read Cyanine Dye (Maleimide) Protein Labeling Protocol]

Cytokine ELISA Protocol - http://www.bdbiosciences.com/pharmingen/protocols/Cytokine_ELISA.shtml

Category: ELISA Protocols: Sandwich ELISA Assay Protocols

Cytokine sandwich ELISA are sensitive enzyme immunoassays that can specifically detect and quantitate the concentration of soluble cytokine and chemokine proteins. BD Biosciences - [Read Cytokine ELISA Protocol]

Cytoskeletal Alterations as a Parameter for Assessment of Toxicity - http://embryo.ib.amwaw.edu.pl/invittox/prot/24.htm#TOP

Category: Pharmacology and Toxicology Protocols: Cytotoxicity Protocols

Changes in the balance of cytoskeletal proteins after exposure to test compounds can be detected by indirect immunofluorescence microscopy and quantitative biochemical methods. - [Read Cytoskeletal Alterations as a Parameter for Assessment of Toxicity]

Deamidation in Proteins and Peptides - http://www.ionsource.com/Card/Deamidation/mono0001.htm

Category: Mass Spectrometry Protocols

Deamidation in Proteins and Peptides. Informational page by Glen Teshima. RPLC, HIC, Peptide mapping of proteolytic digests (LC-MS), Protein carboxyl methyltransferase (PIMT). - [Read Deamidation in Proteins and Peptides]

Denaturing Protein Immunoprecipitation from Mammalian Cells Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4619

Category: Immunology: Immunoprecipitation Protocols: Protein Immunoprecipitation Protocols

Protocol describes a denaturing immunoprecipitation (IP) for mammalian cells. Prefer to use denaturing IPs to recover labeled proteins from pulse-chase experiments. The use of denaturing IPs reduces background considerably. - [Read Denaturing Protein Immunoprecipitation from Mammalian Cells Protocol]

Denaturing Protein Immunoprecipitation from Yeast Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4643

Category: Yeast Protocols: Yeast Protein Protocols: Yeast Immunoprecipitation Protocols

The use of denaturing Ips reduces background and simplifies protein harvest. Yeast are boiled in an SDS-containing solution to liberate total proteins. - [Read Denaturing Protein Immunoprecipitation from Yeast Protocol]

Design, Synthesis, and Characterization of Caged Compounds - http://www.cshprotocols.org/cgi/content/extract/2007/1/pdb.ip25

Category: Microscopy Protocols: Live-Cell Imaging Protocols

When choosing a particular molecule for photoactivation studies, it is necessary to have some structural knowledge of the molecule in order to design an appropriately caged species that will retain its biological inactivity until uncaging is effected. Includes synthesis of caged peptides or proteins. - [Read Design, Synthesis, and Characterization of Caged Compounds]

Detection of Recombinant Proteins by ELISA - http://omrf.ouhsc.edu/~frank/ELISA.html

Category: Recombinant Protein Protocols

ELISA detection of preparation of bacterially expressed protein. ELISA REAGENT RECIPES. Frank Lab. - [Read Detection of Recombinant Proteins by ELISA]

Detection of Recombinant Proteins by ELISA assay - http://omrf.ouhsc.edu/~frank/ELISA.html

Category: ELISA Protocols

Detection of Recombinant Proteins by ELISA protocol. Dr. Frank, Oklahoma Medical Research Foundation. - [Read Detection of Recombinant Proteins by ELISA assay]

Detection of Ubiquitylated Proteins in Yeast Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/30/pdb.prot4615

Category: Yeast Protocols: Yeast Protein Protocols

This assay is performed to detect ubiquitylated proteins in yeast. Yeast that have been transformed with a vector expressing polyhistidine-tagged ubiquitin (Ub) under the control of a copper-inducible promoter are grown, induced with copper, and harvested. Total ubiquitylated proteins are then recovered by nickel-affinity chromatography, and specific proteins are detected by Western blotting. - [Read Detection of Ubiquitylated Proteins in Yeast Protocol]

DNaseI Footprinting Assay - http://www.fhcrc.org/science/labs/breeden/Methods/DNaseI-footprint.html

Category: DNA Protocols: EMSA DNA-Protein Protocols

DNaseI Footprinting Assay. Works well high affinity dna binding proteins. Breeden Lab. - [Read DNaseI Footprinting Assay]

Dried droplet Sample Preparation for MALDI - http://prowl.rockefeller.edu/methods/dried.htm

Category: Mass Spectrometry Protocols

Dried droplet Sample Preparation for MALDI.The discovery of this method allowed the application of laser desorption to proteins [2]. Drying a droplet of a protein/matrix solution remains the favorite method of most MALDI practitioners. Protocol for dried - [Read Dried droplet Sample Preparation for MALDI]

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

Category: Microscopy Protocols: Electron Microscopy Protocols

Article describe the preparation of cells for correlative electron microscopy after live light microscopic observation of fluorescently labeled cytoskeletal proteins microinjected into the same cells. Since identification of cytoskeletal elements in electron microscopic preparations is an essential part of any correlative study, procedures for immunogold labeling of cytoskeletal components and for myosin S1 decoration of actin filaments are also described. - [Read Electron Microscopy of the Cytoskeleton of Cultured Cells]

ENZYME-ASSISTED IMMUNOELECTROBLOTTING (IEB OR WESTERN BLOTTING) - http://www.mcb.uct.ac.za/western.htm

Category: Protein Protocols: Western Blot Protocol

ENZYME-ASSISTED IMMUNOELECTROBLOTTING (IEB OR WESTERN BLOTTING. E.P. Rybicki and Maud Purves. Department of Microbiology. Copper Staining of Gels, Indirect enzyme immunoassay, Staining Proteins on Membranes, Blotting Buffer, Staining of proteins in gels - [Read ENZYME-ASSISTED IMMUNOELECTROBLOTTING (IEB OR WESTERN BLOTTING)]

Epitope Tagging of Recombinant Proteins Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E663998FCCB61070F32F1EAE0301F7F&objectid=6677C749FA75FD435618AC07256968A7

Category: Immunology: Immunoprecipitation Protocols

Protocols for detection and purification of proteins tagged with a particular epitope, the FLAG tag, although the same general approach can be applied to other epitope tags. The protocols employ the anti-FLAG M2 antibody to detect and purify FLAG-tagged proteins. The methods presented are immunoprecipitation of FLAG fusion proteins from cells using an anti-FLAG M2 affinity gel, detection of FLAG fusion proteins by western blotting, and purification of FLAG fusion proteins by anti-FLAG. - [Read Epitope Tagging of Recombinant Proteins Protocol]

EXAMINING THE FUNCTION OF PROTEINS AND PROTEIN NETWORKS WITH THE YEAST TWO-HYBRID SYSTEM - http://proteome.wayne.edu/Update.html

Category: Molecular Model Organisms: Yeast Two-Hybrid Screen Protocol

Interactor Hunt Protocol. Testing whether a bait activates transcription. Screening a library for interactors. Finley Lab. 1997. - [Read EXAMINING THE FUNCTION OF PROTEINS AND PROTEIN NETWORKS WITH THE YEAST TWO-HYBRID SYSTEM]

Experimental Procedures for Protein Microarrays - http://web.archive.org/web/20050313184601/http://cgr.harvard.edu/macbeath/protocols/proteinmicroarrays.html

Category: Protein Microarray Protocols

Experimental Procedures for Protein Microarrays. MacBeath G & Schreiber SL (2000). Printing proteins as microarrays for high-throughput function determination. Science 289: 1760-1763. - [Read Experimental Procedures for Protein Microarrays]

Expression and Purification of His Tagged Proteins in E. coli Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/H1.html

Category: E Coli Protocols: E coli Protein Protocols

Protocol for the expression and purification of His tagged proteins in E. coli. - [Read Expression and Purification of His Tagged Proteins in E. coli Protocol]

Expression of Cloned Genes in E. coli Using IPTG-inducible Promoters Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4085

Category: E Coli Protocols: E coli Protein Protocols

Protocol for the expression of cloned genes in E. coli using IPTG-inducible promoters. Protocol describes how (1) to clone cloned sequences encoding open reading frames in plasmids carrying IPTG-inducible promoters, (2) to optimize expression of target proteins in transformants carrying these recombinants, and (3) to scale-up production of foreign proteins. - [Read Expression of Cloned Genes in E. coli Using IPTG-inducible Promoters Protocol]

Articles (1-3 of 3)

Acid Guanidinium Thiocyanate RNA Isolation Phenol Chloroform Extraction

A single step RNA isolation protocol using Phenol Chloroform Extraction and Acid Guanidinium Thiocyanate. This RNA isolation method uses the fact that guanidinium thiocyanate can simultaneously lyse the cells and inactive cellular RNAses during the initial RNA isolation step allow a single step in the method.

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.