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2-D Polyacrylamide Gel Electrophoresis (Prostate Tissue) - http://cgap-mf.nih.gov/Protocols/DNARNAProteomicAnalysis/Proteomics/2DPage.html

Category: Proteomic Protocols: 2-D Gel Electrophoresis

A detailed protocol for 2-D Polyacrylamide Gel Electrophoresis using Prostate Tissue. Molecular Profiling Initiative. - [Read 2-D Polyacrylamide Gel Electrophoresis (Prostate Tissue)]

2-D Polyacrylamide Gel Electrophoresis Protocol - http://cgap-mf.nih.gov/Protocols/DNARNAProteomicAnalysis/Proteomics/2DPage.html

Category: Primary Cell Isolation and Culture Protocols: Laser Capture Microdissection Protocols

Protocol for 2-D polyacrylamide gel electrophoresis. Includes: 50 ml IEF Lysis Buffer; 10X Electrophoresis Running Buffer (10 L); 30% Acrylamide Stock (1 liter); Separating Acrylamide Gel; 50 ml Equilibration Buffer I; 50 ml Equilibration Buffer II; Transfer Buffer; Sample Preparation; Tissue Processing; Reswelling; Prepare Gradient Acrylamide Gel (9-18%); Transfer and Sequencing of Proteins. - [Read 2-D Polyacrylamide Gel Electrophoresis Protocol]

Detection of DNA in Polyacrylamide Gels by Autoradiography Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4029

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

Detection of radioactive DNA in polyacrylamide gels by autoradiography. - [Read Detection of DNA in Polyacrylamide Gels by Autoradiography Protocol]

Detection of DNA in Polyacrylamide Gels by Staining Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3718

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

Staining bands of DNA in polyacrylamide gels with ethidium bromide, SYBR-Gold, and methylene blue. - [Read Detection of DNA in Polyacrylamide Gels by Staining Protocol]

Gel Retardation Assays for DNA-binding Proteins Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3948

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

Protocol exploits differences in electrophoretic mobility through a nondenaturing polyacrylamide gel between a rapidly migrating target DNA and a more slowly migrating DNA-protein complex. - [Read Gel Retardation Assays for DNA-binding Proteins Protocol]

Histone H1 Kinase Activity Assay - http://www.bio.com/protocolstools/protocol.jhtml?id=p910

Category: Signalling Signal Transduction Protocols

This protocol describes assaying kinase activity of a putative kinase using Histone H1 as the substrate. Histone H1 is the canonical kinase substrate in this type of assay. Phosphorylation of Histone H1 is assessed by SDS-Polyacrylamide gel electrophoresis followed by autoradiography. Includes information: H1 Kinase Assay on Individual Xenopus Oocytes; H1 Kinase Assay on Xenopus Egg Extract Samples; H1 Kinase Assay on Tissue Culture Cells; Helpful protocol hints. - [Read Histone H1 Kinase Activity Assay]

Immunoblot Protocol - http://cgap-mf.nih.gov/Protocols/DNARNAProteomicAnalysis/Proteomics/Immunoblot.html

Category: Primary Cell Isolation and Culture Protocols: Laser Capture Microdissection Protocols

Protocol for Immunoblot. Includes: Staining and Laser Capture Microdissection; Protein Separation by Polyacrylamide Gel Electrophoresis; Electrophoretic Transfer To a Membrane (Nylon, PVDF or Nitrocellulose); Primary and Secondary Antibody Incubations; Visualization. - [Read Immunoblot Protocol]

Immunoblotting: Preparing Protein Solutions Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/21/pdb.prot4298

Category: Western Blot Protocols: General Western Blot Methods Protocols

Protocol describes how proteins are mixed with a sample buffer and prepared for electrophoresis on standard Tris/glycine SDS-polyacrylamide gels. - [Read Immunoblotting: Preparing Protein Solutions Protocol]

Immunoblotting: Semi-Dry Electrophoretic Transfer of Proteins from Gels to Membranes Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4301

Category: Western Blot Protocols: General Western Blot Methods Protocols

The transfer of proteins from a Tris/glycine SDS-polyacrylamide gel to a membrane using a semi-dry method is achieved by placing the gel next to a piece of nitrocellulose filter. This sandwich is placed directly between two plate electrodes, and the proteins are then transferred from the gel onto the filter. - [Read Immunoblotting: Semi-Dry Electrophoretic Transfer of Proteins from Gels to Membranes Protocol]

Immunoblotting: Submerged Electrophoretic Transfer of Proteins from Gels to Membranes Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4302

Category: Western Blot Protocols: General Western Blot Methods Protocols

Transfer of proteins from a Tris/glycine SDS-polyacrylamide gel to a membrane using the submerged method is achieved by placing the gel next to a piece of nitrocellulose filter, submerging this sandwich in a large volume of transfer buffer in a transfer tank, and running current from one side of the transfer tank to another. The proteins are then eluted by transferring them from the gel onto the filter. - [Read Immunoblotting: Submerged Electrophoretic Transfer of Proteins from Gels to Membranes Protocol]

In-Gel Digestion of Proteins Separated by Polyacrylamide Gel Electrophoresis - http://web.archive.org/web/20010906052645/http://www.pharma.ethz.ch/bmm/protocols/ingeldig.html

Category: Proteomic Protocols: 2-D Gel Electrophoresis

In gel trypsin digestion for Mass spectrometry. (According to EMBL Protein and Peptide Group, 1997) - [Read In-Gel Digestion of Proteins Separated by Polyacrylamide Gel Electrophoresis]

In-gel digestion of proteins to be analyzed by mass spectrometry - http://www.narrador.embl-heidelberg.de/GroupPages/PageLink/activities/protocols/ingeldigest.html

Category: Proteomic Protocols

In-Gel Digestion Protocol. Excision of protein bands (spots) from polyacrylamide gels Reduction and alkylation . Reduction and alkylation . MALDI analysis of the supernatant after in-gel digestion. Extraction of Peptides.Matthias Wilm EMBL Bioanalytical R - [Read In-gel digestion of proteins to be analyzed by mass spectrometry]

In-Gel Kinase Assay - http://www.bio.com/protocolstools/protocol.jhtml?id=p911

Category: Signalling Signal Transduction Protocols

Protocol describes how to assay for kinase activity within a polyacrylamide gel, rather than in solution. The advantages to an in-gel assay are that an apparent molecular weight can be assigned to the kinase activity and that multiple kinase activities can be distinguished. Includes protocol hints. - [Read In-Gel Kinase Assay]

Isolation of DNA Fragments from Polyacrylamide Gels by the Crush and Soak Method Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot2936

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

The "crush and soak" method, which works best for DNAs <1 kb in size, can be used to recover both singleand double-stranded DNAs from polyacrylamide gels. The yield of eluted DNA varies from <30% to >90% depending on the size of the DNA fragment. - [Read Isolation of DNA Fragments from Polyacrylamide Gels by the Crush and Soak Method Protocol]

Kinase Assay Using In Vitro Translated Xenopus Mos Kinase - http://www.bio.com/protocolstools/protocol.jhtml?id=p912

Category: Signalling Signal Transduction Protocols

Protocol describes how tagged Mos kinase is translated in vitro, immunopurified, and used in a kinase assay. Protocol includes: Translation of Xenopus Mos Kinase; Antibody to Antigen Binding; Protein A Sepharose to Antibody Binding; Kinase Reactions on Immunoprecipitated Material; Polyacrylamide Gel Analysis of Immunoprecipitates. Includes protocol hints. - [Read Kinase Assay Using In Vitro Translated Xenopus Mos Kinase]

Neutral Polyacrylamide Gel Electrophoresis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4028

Category: Nucleic Acid Electrophoresis Protocols

Protocol on how to pour and run a neutral polyacrylamide gel. - [Read Neutral Polyacrylamide Gel Electrophoresis Protocol]

Preparation of Denaturing Polyacrylamide Gels Containing Formamide Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3807

Category: DNA Protocols: DNA Sequencing Protocols

The inclusion of formamide in sequencing gels eliminates secondarystructure in the DNA during electrophoresis. Formamide gelsare particularly useful and almost a necessity when sequencingDNA templates with a G/C content >55%. - [Read Preparation of Denaturing Polyacrylamide Gels Containing Formamide Protocol]

Preparation of Denaturing Polyacrylamide Gels Containing Formamide Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3807

Category: Nucleic Acid Electrophoresis Protocols

Protocol for preparation of denaturing polyacrylamide gels containing formamide.The inclusion of formamide in sequencing gels eliminates secondary structure in the DNA during electrophoresis. Formamide gels are particularly useful and almost a necessity when sequencing DNA templates with a G/C content >55%. - [Read Preparation of Denaturing Polyacrylamide Gels Containing Formamide Protocol]

Preparation of Denaturing Polyacrylamide Gels Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3793

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

Protocol for preparation of denaturing polyacrylamide gels. - [Read Preparation of Denaturing Polyacrylamide Gels Protocol]

Preparation of Polyclonal Antibodies from Plasma or Serum Using the Gradiflow BF400 - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4236

Category: Antibody Protocols: Antibody Production Protocols

Polyclonal antibodies can be isolated from animal plasma or serum using the procedure described in this protocol. The Gradiflow BF400 instrument has two liquid streams that circulate through a separation cartridge positioned between two electrodes and composed of three hydrogel polyacrylamide membranes, which define the channels for the two sample streams. The central membrane forms a physical barrier between the two streams. - [Read Preparation of Polyclonal Antibodies from Plasma or Serum Using the Gradiflow BF400]

Principles and Applications of Liquid Scintillation Counting PDF - http://www.ehs.psu.edu/radprot/LSC_Theory2.pdf

Category: Radioactivity: Liquid Scintillation Counting

Excellent guide for Liquid Scintillation Counting. Includes protocols and methods for counting gel slices, SPECIAL SAMPLE PREPARATION PROTOCOLS: TLC Plates, protocol for Counting Samples on Cellulose-ester, Filters (MilliporeTM filters), Counting Tissue radioacitivity, Counting 14CO2, Samples in Polyacrylamide Gels. National Diagnostics. National Diagnostics Laboratory Staff. - [Read Principles and Applications of Liquid Scintillation Counting PDF]

Protocol for Polymerase III In Vitro Transcription - http://www.bio.com/protocolstools/protocol.jhtml?id=p1140

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

Protocol describes an RNA Polymerase III (Pol III) transcription assay using an extract or proteins of choice. Pol III is the polymerase responsible for transcribing 5S RNA, tRNAs, and other small RNAs. Î±-Amanitin inhibits Pol II transcription in the assay. The newly-transcribed, radiolabeled RNA is visualized by autoradiography following Urea Polyacrylamide gel electrophoresis. - [Read Protocol for Polymerase III In Vitro Transcription]

Purification of DNA Recovered Anion-exchange Chromatography Protocol - http://cshprotocols.cshlp.org/cgi/content/extract/2006/2/pdb.prot4024

Category: Chromatography Protocols: Ion Exchange Chromatography Protocols

Protocol for the purification of DNA recovered from agarose and polyacrylamide gels by anion-exchange chromatography. Fragments of DNA recovered from agarose gels are sometimes poor templates or substrates in subsequent enzymatic reactions. This problem can be solved by binding the DNA to a positively charged matrix, such as DEAE-Sephadex or DEAE-Sephacel, in buffers of low ionic strength. After washing the matrix, the DNA is eluted by raising the strength of the buffer. - [Read Purification of DNA Recovered Anion-exchange Chromatography Protocol]

Purification of Synthetic Oligonucleotides by Polyacrylamide Gel Electrophoresis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4063

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

Protocol for purification of synthetic oligonucleotides by polyacrylamide gel electrophoresis. As a rule of thumb, oligonucleotides >25 nucleotides should be purified by polyacrylamide gel electrophoresis, as should oligonucleotides of any length that yield anomalous results. After electrophoresis, the oligonucleotide is eluted from the gel and concentrated by reversed-phase chromatography on Sep-Pak C18 columns. - [Read Purification of Synthetic Oligonucleotides by Polyacrylamide Gel Electrophoresis Protocol]

Recovery of DNA from Agarose and Polyacrylamide Gels: Electroelution into Dialysis Bags Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4023

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Agarose Gels Protocols

A messy but reliable technique that works well for DNAs ranging in size from 200 bp to >50 kb. - [Read Recovery of DNA from Agarose and Polyacrylamide Gels: Electroelution into Dialysis Bags Protocol]

Articles (1-4 of 4)

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.

DNA Fragment Purification from Polyacrylamide Gels Protocol

A simple protocol for the purification of DNA fragments from Polyacrylamide Gels.

Histone H1 Kinase Activity Assay

Histone H1 Kinase Activity Assay Protocol. This protocol describes assaying kinase activity of a putative kinase using Histone H1 as the substrate. Histone H1 is the canonical kinase substrate in this type of assay. Phosphorylation of Histone H1 is assessed by SDS-Polyacrylamide gel electrophoresis followed by autoradiography.