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Lysing Tissue-Culture Cells for Immunoprecipitation Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/23/pdb.prot4531

Category: Cell Culture Protocols: Cell Lysis Protocols

For cells grown in tissue culture, the most useful method of lysis is treating with detergents, as described in this protocol. Non-ionic detergents, such as NP-40, solubilize the plasma and intracellular membranes, break many weak intermolecular bonds, and solubilize most of the commonly studied protein antigens. RIPA lysis buffer may be used as a more rigorous extraction buffer to release all but the insoluble proteins of the cell and to break most weak noncovalent interactions. - [Read Lysing Tissue-Culture Cells for Immunoprecipitation Protocol]

Lysis and Protein Extraction from WEHI-231 Cells with TriPure Isolation Reagent Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000162.pdf?pid=PP00000162

Category: Chromatography Protocols: Immobilized Metal Ion Affinity Chromatography Protocols

Protocol is the first step in a three-step process for the preparation and enrichment of phosphopeptides using immobilized metal affinity chromatography (IMAC) for the identification of the phosphopeptides by LC-MS/MS. This procedure is used to prepare protein extracts from WEHI-231 cells. This preparation method provides total cellular protein samples that are free of contaminating nucleic acids. - [Read Lysis and Protein Extraction from WEHI-231 Cells with TriPure Isolation Reagent Protocol]

Mini-Prep Protocol for Purification of Plasmid DNA from Bacteria - http://people.morehead-st.edu/fs/d.peyton/protocols.html

Category: Microbiology: Mini-Prep Plasmid Purification

Using Solutions A for Lysis, Solutions B for Neutralization Solution, and Solution C for washing. Dr.David Peyton. - [Read Mini-Prep Protocol for Purification of Plasmid DNA from Bacteria]

Non-denaturing Cell Lysis Buffer - http://www.chemicon.com/techsupp/celllysis.asp

Category: Buffers Media, and Solutions: Buffer Recipes for Protein Work

Non-denaturing Cell Lysis Buffer. Chemicon. - [Read Non-denaturing Cell Lysis Buffer]

Preparation of cell lysates from yeast using glass beads vortexing - http://www.embl.de/ExternalInfo/protein_unit/draft_frames/protocol_database/Purification/ec_glass_beads.html

Category: Protein Protocols: Protein Extraction From Cells

Preparation of cell lysates from yeast using glass beads vortexing. Includes lysis buffer recipes. The Protein Expression and Purification Facility, EMBL - [Read Preparation of cell lysates from yeast using glass beads vortexing]

Preparation of DNA for Pulsed-field Gel Electrophoresis: Isolation of Intact DNA from Yeast - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3235

Category: Nucleic Acid Electrophoresis Protocols: Pulsed Field Gel Electrophoresis Protocols

Protocol for preparation of DNA for pulsed-field gel electrophoresis: isolation of intact DNA from yeast. Yeast cells are first treated enzymatically to break down the cell walls and then resuspended in low-melting-temperature agarose plugs. The DNA is liberated by infusing the plugs with lysis buffer and proteases. This method is used to prepare both conventional and artificial yeast chromosomes. - [Read Preparation of DNA for Pulsed-field Gel Electrophoresis: Isolation of Intact DNA from Yeast]

Preparation of Plasmid DNA by Alkaline Lysis with SDS: Maxipreparation Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4090

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from large-scale (500 ml) bacterial cultures by treatment with alkali and SDS. - [Read Preparation of Plasmid DNA by Alkaline Lysis with SDS: Maxipreparation Protocol]

Preparation of Plasmid DNA by Alkaline Lysis with SDS: Midipreparation Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3901

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from intermediate-scale (20-50 ml) bacterial cultures by treatment with alkali and SDS. - [Read Preparation of Plasmid DNA by Alkaline Lysis with SDS: Midipreparation Protocol]

Preparation of Plasmid DNA by Alkaline Lysis with SDS: Minipreparation Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4084

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from small-scale (1-2 ml) bacterial cultures by treatment with alkali and SDS. - [Read Preparation of Plasmid DNA by Alkaline Lysis with SDS: Minipreparation Protocol]

Preparation of Plasmid DNA by Large-scale Boiling Lysis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3910

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from large-scale (500 ml) bacterial cultures by treatment with Triton X-100 and lysozyme, followed by heating. This method is not recommended for preparing plasmid DNA from strains of E. coli that express endonuclease A (endA+ strains). - [Read Preparation of Plasmid DNA by Large-scale Boiling Lysis Protocol]

Preparation of Plasmid DNA by Lysis with SDS Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3906

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Large (>15 kb), closed circular plasmids are prepared (albeit inefficiently and in small yield) by lysing bacteria with SDS. - [Read Preparation of Plasmid DNA by Lysis with SDS Protocol]

Preparation of Plasmid DNA by Small-scale Boiling Lysis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3903

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from small-scale (1-2 ml) bacterial cultures by treatment with Triton X-100 and lysozyme, followed by heating. This method is not recommended for preparing plasmid DNA from strains of E. coli that express endonuclease A (endA+ strains). - [Read Preparation of Plasmid DNA by Small-scale Boiling Lysis Protocol]

Preparation of RNA from Plant Tissue Using Guanidinium Isothiocyanate/Cesium Chloride Ultracentrifug - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4102

Category: Plant Biology Protocols: Plant DNA RNA Isolation Protocols

Tissues are homogenized in guanidinium isothiocyanate lysis buffer and then subjected to cesium chloride ultracentrifugation. The resulting RNA is then extracted with phenol and chloroform to remove protein and lipid contaminants. - [Read Preparation of RNA from Plant Tissue Using Guanidinium Isothiocyanate/Cesium Chloride Ultracentrifug]

Preparing Stocks of Bacteriophage {lambda} by Plate Lysis and Elution Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3963

Category: Viral Culture and Isolation Protocols: Viral Culture Protocols

A reliable method to prepare plate lysates and to recover infectious bacteriophages by elution from the top agar. - [Read Preparing Stocks of Bacteriophage {lambda} by Plate Lysis and Elution Protocol]

Protocol for Competitive Ligand Binding to Cortical Receptor using Crosslinking - http://www.bio.com/protocolstools/protocol.jhtml?id=p1254

Category: Signalling Signal Transduction Protocols

Protocol describes a competitive ligand binding assay for cortical neurotrophin receptors. Following binding in the presence of competitor, the bound radiolabeled ligand is cross-linked to the receptor. The cells are lysed and the ligand-receptor complexes are immunoprecipitated using a pan-trk (tyrosine kinase receptor) antibody. Protocol includes:Preparation of Cortical Tissue for Competitive Crosslinking, Competitive Binding, Crosslinking Ligand to Receptor, Lysis and Immunoprecipitation etc - [Read Protocol for Competitive Ligand Binding to Cortical Receptor using Crosslinking]

Protocol for Immunopurification of Tyrosine Phosphorylated Proteins - http://www.bdbiosciences.com/pharmingen/protocols/Tyrosine_Phosphorylated_Proteins.shtml

Category: Signalling Signal Transduction Protocols

Protocol for immunopurification of tyrosine phosphorylated proteins. Protocol includes: Cell Lysis (Non-Denaturing and Denaturing Conditions) and Purification. - [Read Protocol for Immunopurification of Tyrosine Phosphorylated Proteins]

Protocol for JNK Kinase Assay - http://www.whitelabs.org/Lab%20Protocols/kinase%20and%20phosphatase%20assays/JNK%20Kinase%20Assay.htm

Category: Signalling Signal Transduction Protocols: MAPK Protocols

Protocol describing a JNK kinase asssay. Protocol includes information on: Triton Lysis Buffer, Kinase Assay Buffer, Kinase Assay and Preparation of kinase - [Read Protocol for JNK Kinase Assay]

Protocol Immunoprecipitation - http://www.sigmaaldrich.com/Area_of_Interest/Life_Science/Antibody_Explorer/Procedures/Immunoprecipitation.html

Category: Immunology: Immunoprecipitation Protocols

Immunoprecopitation method, the protein from the cell or tissue homogenate is precipitated in an appropriate lysis buffer by means of an immune complex which includes the antigen (protein), primary antibody and Protein A-, G-, or L-agarose conjugate or a secondary antibody-agarose conjugate - [Read Protocol Immunoprecipitation]

Quick DNA Plasmid Prep Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/D4.html

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Protocol for quick DNA plasmid prep. Protocol gives very clean plasmid preps for restriction digests and cloning. However, due to the alkaline lysis step, the DNA is often nicked and may not give exceptional sequence data. - [Read Quick DNA Plasmid Prep Protocol]

Raf-1 Kinase Assay - http://www.bio.com/protocolstools/protocol.jhtml?id=p1980

Category: Signalling Signal Transduction Protocols: MAPK Protocols

This protocol uses mitogen-activated protein/ERK kinase (MEK) to activate the extracellular-signal-regulated (Erk) mitogen-activated protein (MAP) kinases upon agonist binding to receptors. Protocol includes information about: Harvest, Immunoprecipitation and Kinase Reaction. Also includes following solutions: Laemmli Sample Buffer (1X), ATP Mix, Kinase Buffer (10X), Lysis Buffer. Helpful protocol hints are also included. - [Read Raf-1 Kinase Assay]

Tetracycline as Regulator of Inducible Gene Expression III - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3958

Category: Transfection Protocols: Stable Transfection Protocols

Stably transfected cells, generated in the first two stages of the procedure, are induced for expression of the target gene. After harvesting and lysis, the lysates are analyzed by SDS-PAGE and immunoblotting. - [Read Tetracycline as Regulator of Inducible Gene Expression III]

Ultimate Freeze Thaw Lysis For Mammalian Cells - http://tanseylab.cshl.edu/pdf/Freeze_Thaw_Lysis.pdf

Category: Protein Protocols: Protein Extraction From Cells

Ultimate Freeze Thaw Lysis For Mammalian Cells. Tansey lab. Cell harvesting. Includes recipes for solutions for freeze thawing lysis for cells. - [Read Ultimate Freeze Thaw Lysis For Mammalian Cells]

Western Blotting and Recipies - http://www.ualberta.ca/PERINATAL/facilities/pdf/Western--Protein%20preparation.pdf

Category: Buffers Media, and Solutions: Buffer Recipes for Protein Work

Western Blotting and Recipies. Lysis Buffer. - [Read Western Blotting and Recipies]

Yeast Plasma Membrane H+ -ATPASE Toxcity Test Protocol - http://embryo.ib.amwaw.edu.pl/invittox/prot/34.htm

Category: Yeast Protocols

Plasma membranes are isolated from the yeast Saccharomyces cerevisiae. The cell wall is initially digested by helicase, followed by hypoosmotic lysis and homogenization. Membranes are prepared by subsequent differential centrifugation. The activity of the H+-ATPase is then determined by measuring the amount of inorganic phosphate released from ATP. - [Read Yeast Plasma Membrane H+ -ATPASE Toxcity Test Protocol]

Articles (1-10 of 11)

Populational Analysis of Genomic DNA Protocol

A protocol on the populational Analysis of genomic DNA.

Lambda Phage Protocol for Large Preparation DNA Recovery

Lambda Phage Protocol for Large Preparations DNA Recovery.

Lambda Phage Protocol Large DNA Preparation

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.

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.

Toluidine Blue Stain

Toluidine Blue staining method and protocol for cell or tissue histology.

Thionin Stain

Thionin staining method and protocol for cell or tissue histology.

Picking Transfected ES Cell Clones Protocol

This protocol a protocol on how to generate transfected embryonic stem (ES) cell clones. The previous protocol in this series is the Protocol for Electroporation of ES cells. The next protocol in the series is the Protocol on Disaggregation, Expansion, and Freezing of Transfected ES Clones.

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