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Actin Staining in Fixed Yeast Cells Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4168

Category: Cytoskeleton Protocols: Actin Myosin Protocols

Phalloidin binds specifically to F-actin, and fluorescent-tagged phalloidin stains the actin skeleton in cells in a manner that is very close to the staining pattern seen using anti-actin antibody. - [Read Actin Staining in Fixed Yeast Cells Protocol]

Anti Peroxidase Technique - http://www.nottingham.ac.uk/pathology/protocols/papihc.html

Category: Immunohistochemistry Protocols

Anti peroxidase technique. Includes: DAB Solution. - [Read Anti Peroxidase Technique]

Anti-Nogo-A Treatment Protocol - http://www.natureprotocols.com/2006/07/21/antinogoa_treatment_enhances_c.php

Category: Neuroscience Protocols

After section of the corticospinal (CS) tract in adult rats, neutralizing Nogo-A with monoclonal antibodies leads to enhancement of axonal re-growth and compensatory sprouting, accompanied by increased motor recovery. The neutralization of Nogo-A represents a promising approach for therapy after lesion if its enhancement of functional recovery can be transposed to primates. - [Read Anti-Nogo-A Treatment Protocol]

Binding Antibodies to Tissue Sections Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4331

Category: Antibody Protocols: Antibody Labeling Protocols

Once tissues are fixed and permeabilized, the antibodies are added. These antibodies can be labeled directly or detected by a labeled secondary reagent. For indirect detection, any reagent that binds specifically to the primary antibody can be "tagged" and used to locate the antibody. The possible reagents include anti-immunoglobulin antibodies, protein A or G, or, if the first antibody is labeled with biotin, streptavidin. They can be labeled with enzymes or gold. - [Read Binding Antibodies to Tissue Sections Protocol]

CHO Centrosome Prep Protocol - http://mitchison.med.harvard.edu/protocols/mt1.html

Category: Cell Organelle Protocols: Nucleus Protocols

The key step is the lysis which solubilizes centrosomes away from nuclei by very low ionic strength lysis after treatment of cells with nocodazole and cytochalasin B. The released centrosomes are then centrifuged onto a Ficoll cushion (to avoid pelleting) and the interface between the lysate and the Ficoll is collected and the centrosomes are concentrated on a sucrose gradient. Fractions are assayed by spindown and double IF with 5051 serum and anti-tubulin and the pooled fractions are frozen... - [Read CHO Centrosome Prep Protocol]

Detection FISH Protocol - http://www.riedlab.nci.nih.gov/publications/FISH%20Detection.pdf

Category: Cytogenetics Protocols: Fluorescent In Situ Hybridization FISH Protocol

Protocol for Detection (FISH). TRITC, or Avidin Cy-5. For the probes labeled with digoxigenin, we usually first incubate with mouse-anti-digoxigenin, followed by incubation with sheep anti-mouse Cy5.5, or other fluorochrome conjugated antibodies. - [Read Detection FISH Protocol]

ELISA for detection of platelet-associated Ig (PAIg) - http://www.icp.ucl.ac.be/mexp/vipg_plelisa.html

Category: ELISA Protocols

Qualitative ELISA (Enzyme-Linked Immunosorbent Assay) used for screening detection of anti-platelet antibodies or for detection of platelet-associated Ig (PAIg). Andrei Musaji Viral immunity and pathogenesis group, Universite Catholique de Louvain. - [Read ELISA for detection of platelet-associated Ig (PAIg)]

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]

Erythrophagocytosis Assay - http://www.icp.ucl.ac.be/mexp/vipg_eph.html

Category: Immunology: Macrophage Monocyte Protocols

Erythrophagocytosis Assay. The erythrophagocytosis assay is performed to compare the phagocytic rate with and without anti-eythrocyte antibody in either macrophages from control animals or virus-infected counterparts. Andrei Musaji Viral Immunity and Pathogenesis Group - [Read Erythrophagocytosis Assay]

Fluorescent In Situ Transcription in Cells and Tissues Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E6639A7FEDD7A49BBC3758516F99557&objectid=6677C9E8B913BFB72E9F654025F3C39A

Category: Cytogenetics Protocols: Fluorescent In Situ Hybridization FISH Protocol

Method assesses cellular mRNA transcripts in tissue sections and cell cultures using unique short anti-sense primers directed against sequences in particular protein(s). The unlabeled synthetic cDNA oligonucleotide primers are extended complementary to a sense mRNA transcript using reverse transcriptase and labeled through incorporation of a fluorescent-labeled dUTP nucleotide base. This procedure provides rapid detection of low abundance mRNA messages that can be related to other cellular.... - [Read Fluorescent In Situ Transcription in Cells and Tissues Protocol]

Human T Cell Activation Protocol - http://www.ebioscience.com/ebioscience/appls/AC145.htm#human

Category: Immunology: T Cell Protocols

Protocol for the Stimulation of human peripheral blood mononuclear cells with anti-human CD3 monoclonal antibody; MTT assay for detection of cellular proliferation. Human PBMCs can be activated in vitro by soluble anti-human CD3 antibodies. Performed titration studies with these antibodies and established the following protocol for stimulation of PBMC. - [Read Human T Cell Activation Protocol]

Identification of Single Bacterial Cells using DIG-Labeled Oligonucleotides Protocol - http://www.roche-applied-science.com/PROD_INF/MANUALS/InSitu/pdf/ISH_139-142.pdf

Category: Cytogenetics Protocols: DIG Biotin Labeled Probe Protocols

Protocol for the identification of single bacterial cells using DIG-labeled oligonucleotides. Includes: Organisms and growth conditions; Cell fixation and preparation of cell smears; DIG labeling of oligonucleotides with DIG-ddUTP; In situ hybridization using digoxigenin-labeled oligonucleotides; Detection of DIG-labeled oligonucleotides with fluorescently labeled anti-DIG Fab fragments; Detection of DIG-labeled oligonucleotide. - [Read Identification of Single Bacterial Cells using DIG-Labeled Oligonucleotides Protocol]

Immunoblotting: Antigen Detection Using Chemiluminescence Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4271

Category: Western Blot Protocols: Antigen Detection Methods Protocols

The blot is blocked to prevent nonspecific adsorption of the immunological reagents. Antibodies are then bound to the proteins immobilized on the membrane, and the antigen is detected by labeling the antibodies with conveniently identified tags. Common labeling methods for chemiluminescent detection include anti-immunoglobulin antibody-coupled enzymes such as horseradish peroxidase, which catalyzes the oxidation of luminol and in turn releases light. - [Read Immunoblotting: Antigen Detection Using Chemiluminescence Protocol]

Immunocytochemical Identification of Proliferating Cells In Vascular Tissue - http://www.emory.edu/WILCOX/brduDAB.html

Category: Immunology: Immunocytochemistry Protocols

Using anti-BrdU, Diaminobenzadine (DAB) brown detection. Includes counterstaining protocol. - [Read Immunocytochemical Identification of Proliferating Cells In Vascular Tissue]

Immunofluorescence using Anti-Serotonin Antibody in C. elegans Protocol - http://www.bio.com/protocolstools/protocol.jhtml?id=p1561

Category: C. elegans Protocols

Protocol for immunofluorescence using anti-serotonin antibody in C. elegans. - [Read Immunofluorescence using Anti-Serotonin Antibody in C. elegans Protocol]

Immunofluorescent Staining of Intracellular Cytokines for Flow Cytometric Analysis - http://www.bdbiosciences.com/pharmingen/protocols/Intracellular_Cytokines.shtml

Category: Signalling Signal Transduction Protocols: Cytokine Protocols

Fluorescent anti-cytokine and monoclonal antibodies are useful for the intracellular staining and multiparameter flow cytometric analysis of individual cytokine-producing cells within mixed cell populations. BD Biosciences. - [Read Immunofluorescent Staining of Intracellular Cytokines for Flow Cytometric Analysis]

Immunohistochemistry using Anti-Ganglioside Antibodies - http://www.glycotech.com/protocols/Proto2.html

Category: Molecular Biology Protocols: Carbohydrate Protocols

Indirect method measuring immunofluorescence coupled to second antibody. Best for membrane antigens in addition to intra- and extracellular antigens, may be applied to frozen tissue sections, to cells in suspension, and to cells attached to glass slides or coverslips. Tadashi Tai~Head, Department of Tumor Immunology, The Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan - [Read Immunohistochemistry using Anti-Ganglioside Antibodies]

Immunoprecipitation with Anti-Phosphotyrosine:Biotin Conjugates Protocol - http://www.bdbiosciences.com/pharmingen/protocols/Biotin_Conjugates.shtml

Category: Immunology: Immunoprecipitation Protocols

Protocol for immunoprecipitation with anti-phosphotyrosine:biotin conjugates. Includes: Denaturing Conditions; Native Conditions; Pre-clearing. - [Read Immunoprecipitation with Anti-Phosphotyrosine:Biotin Conjugates Protocol]

Immunoscreening of Lambda-GT11 Bacteriophage Libraries with Antibody Probes - http://omrf.ouhsc.edu/~frank/GT11.html

Category: RNA Protocols: cDNA Libraries Library

Adsroption of Anti-E. coli Antibodies, cDNA Library Screening, Frank Lab. Oklahoma Medical Research Foundation - [Read Immunoscreening of Lambda-GT11 Bacteriophage Libraries with Antibody Probes]

Improved Detection of Apoptotic Cells in Archival Paraffin Sections: Immunohistochemistry Using Anti - http://www.jhc.org/cgi/content/full/50/4/449

Category: Cell Biology and Cell Culture: Apoptosis Protocols

Improved Detection of Apoptotic Cells in Archival Paraffin Sections: Immunohistochemistry Using Antibodies to Cleaved Caspase 3. Gown et al, 2002 - [Read Improved Detection of Apoptotic Cells in Archival Paraffin Sections: Immunohistochemistry Using Anti]

Isolation of Resting B Lymphocytes from One or More Groups of Four Mouse Spleens Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000001.pdf?pid=PP00000001

Category: Immunology: B Cell Protocols

Describes the isolation of resting B lymphocytes (B cells) from mouse spleens by using negative selection with anti-CD43 and anti-Mac-1/CD11b monoclonal antibodies coupled to magnetic microbeads. This strategy depletes non-B cells from a mixed population of splenocytes and relies on the fact that most mature leukocytes, with the exception of resting splenic B cells, express CD43. - [Read Isolation of Resting B Lymphocytes from One or More Groups of Four Mouse Spleens Protocol]

Isolation of Resting B Lymphocytes from Sixteen Mouse Spleens Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000016.pdf?pid=PP00000016

Category: Immunology: Mononuclear Cell Isolation Protocols: B Lymphocyte Isolation Protocols

Protocol describes the isolation of resting B lymphocytes (B cells) from mouse spleens by using negative selection with anti-CD43 and anti-Mac-1/CD11b monoclonal antibodies coupled to magnetic microbeads. This strategy depletes non-B cells from a mixed population of splenocytes and relies on the fact that most mature leukocytes, with the exception of resting splenic B cells, express CD43. - [Read Isolation of Resting B Lymphocytes from Sixteen Mouse Spleens Protocol]

Isolation of Resting B Lymphocytes from Sixteen Mouse Spleens Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000016.pdf?pid=PP00000016

Category: Immunology: B Cell Protocols

Isolation of Resting B Lymphocytes Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000001.pdf?pid=PP00000001

Category: Immunology: Mononuclear Cell Isolation Protocols: B Lymphocyte Isolation Protocols

This procedure describes the isolation of resting B lymphocytes (B cells) from mouse spleens by using negative selection with anti-CD43 and anti-Mac-1/CD11b monoclonal antibodies coupled to magnetic microbeads. This strategy depletes non-B cells from a mixed population of splenocytes and relies on the fact that most mature leukocytes, with the exception of resting splenic B cells, express CD43. - [Read Isolation of Resting B Lymphocytes Protocol]

Protocol for Antiphosphotyrosine Western Blot Analysis - http://www.bio.com/protocolstools/protocol.jhtml?id=p697

Category: Signalling Signal Transduction Protocols

Detection of phosphorylated tyrosine residues can be performed using anti-P-TYR Ab and Western Analysis.Includes 2nd method,which uses phosphotyrosine in conjunction with anti-P-TYR Ab to "unlabel" potential proteins.By comparing Westerns developed with the 1st method(reveals phosphorylated protein) and the 2nd method(reveals non-specific labeling), a more accurate picture of those proteins phosphorylated on tyrosine can be seen. Includes: Protein Preparation, Electrophoresis and Transfer. - [Read Protocol for Antiphosphotyrosine Western Blot Analysis]

Articles (1-9 of 9)

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.

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.

Coupling Cysteine Containing Peptides to Carrier Protein for Immunization and Polyclonal Antibody

This protocol is used in the production of polyclonal antibodies that recognize a peptide sequence of interest.

Absorption Spectroscopy and Quantification of Filamentous Phage Protocol

Unlike spherical phage, such as T4 and λ, which have roughly equal weight ratios of protein to DNA, filamentous phage have about six times more protein than DNA; the protein therefore contributes substantially to the absorption spectrum.

3' Rapid Amplification of cDNA Ends RACE Using PCR Protocol

3' Rapid Amplification of cDNA Ends RACE Using PCR Protocol. This protocol contains the steps for 3' end rapid amplification of mRNA by PCR. The first-strand cDNA is synthesized from total or poly(A+) RNA by priming from the poly-A tail of the mRNA using a oligo (dT) adaptor primer. The cDNA is then amplified via PCR using a gene-specific primer and an adaptor primer.

Electrotransformation of BMH 81-17mut S for Isolating Site-Directed dsDNA Mutants

This protocol describes the electroporation of the BMH 81-17 mut S strain that is recommended for tranformation of the site directed mutagenesis of dsDNA (See Protocol on Site-Directed Mutagenesis on Double Stranded DNA). BMH 81-17 mut S are a mismatch repair defective (mut S) Escherichia coli strain. The probability that the two mutations will cosegregate during the first round of DNA replication is increased in this strain.

Vector Design Protocol for Transgenic Mice Generation

The protocol gives general considerations for the design of targeting vectors for transgenic mice. The protocol shares tips in the design of knock-out and knock-in vectors and some of their strategies for producing homologously recombined embryonic stem cells.

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