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Western Blot Detailed (PDF) - http://www.its.caltech.edu/%7Ebjorker/Western_blot.pdf

Category: Protein Protocols: Western Blot Protocol

Protocol includes cell lysate preparation, SDS-PAGE gels, transfer, and blotting. (Bjorkman Group). - [Read Western Blot Detailed (PDF)]

TAIL PCR Protocol - http://preuss.bsd.uchicago.edu/protocols/tail.html

Category: PCR Protocols

TAIL PCR Protocol. TAIL is a series of reactions that are intended to map where a T-DNA (transfer DNA) has inserted within the genome. The main components of the 3 reactions are the AD (Arbitrary Degenerate) primers, border primers, and DNA from the T-DNA - [Read TAIL PCR Protocol]

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]

Western Blot Troubleshooting - http://www.westernblotinfo.com/western-blot-troubleshooting/

Category: Protein Protocols: Western Blot Protocol: Troubleshooting Western Blots

Streaking of Western Blots, Weak Bands or Weak Staining of Western Blots, Poor Transfer of Membranes, Dirty Blot, Multiple Bands on Western Blot Membrane and Film. Western Blot Info. - [Read Western Blot Troubleshooting]

Troubleshooting Guide: Western Blot - http://www.rndsystems.com/tsg_detail_objectname_western_blot.aspx

Category: Protein Protocols: Western Blot Protocol: Troubleshooting Western Blots

Problem: Poor Transfer, Problem: Poor Transfer. R&D Systems. - [Read Troubleshooting Guide: Western Blot]

Intracellular Fluorescent Dye CFSE to Monitor Lymphocyte Migration and Proliferation - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E6634D5A5D8B91EC631DE5426302733&objectid=6674D9B6A53E5856FBF23C4E94630203

Category: Immunology: T Cell Protocols

Describes methods for labeling high or low numbers of lymphocytes with CSFE. Protocols are provided to use CSFE-labeled cells in cell transfer studies or as cells to be cultured in vitro. Detailed guidelines for positioning of CSFE-labeled lymphocytes in lymphoid organs or other tissues are included for those wishing to use this approach to study lymphocyte migration. - [Read Intracellular Fluorescent Dye CFSE to Monitor Lymphocyte Migration and Proliferation]

Staining Immunoblots for Total Protein Using Ponceau S Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4269

Category: Western Blot Protocols: Immunoblot Stains for Total Protein Protocols

Protocol used to determine the position of molecular-weight markers and to ensure that efficient transfer of proteins to the blot has occurred, the total composition of the proteins can be determined by staining the membrane with Ponceau S . - [Read Staining Immunoblots for Total Protein Using Ponceau S Protocol]

Western Blot Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/W1.html

Category: Western Blot Protocols: General Western Blot Methods Protocols

This protocol was developed for the BIORAD protein gel and transfer apparatus. The buffers can be used with any electrophoresis/transfer system. - [Read Western Blot Protocol]

Optimizing Western Blots: Pre-Transfer Invitrogen - http://www.invitrogen.com/content.cfm?pageid=10713

Category: Protein Protocols: Western Blot Protocol: Troubleshooting Western Blots

Optimizing Western Blots: Pre-Transfer Invitrogen - [Read Optimizing Western Blots: Pre-Transfer Invitrogen]

Blastocyst Embryo Transfer into Pseudopregnant Mouse Protocol - http://members.cox.net/microinjectionworkshop/technical/tnp6.html

Category: Mouse Protocols: Mouse Embryology Protocols

Protocol for blastocyst embryo transfer into pseudopregnant recipient female mouse. - [Read Blastocyst Embryo Transfer into Pseudopregnant Mouse Protocol]

Preparation and LCM of Paraffin Embedded Tissue Sections Protocol - http://dir.nichd.nih.gov/lcm/LCMTAP.htm#II.%20Preparation%20and%20LCM%20of%20Paraffin%20Embedded%20Tissue%20Sections

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

Protocol for the preparation of LCM of Paraffin embedded tissue sections. Includes: Fixation; Processing, Embedding and Tissue Sectioning; Staining; Laser Transfer; DNA Extraction Protocol of LCM tissue and PCR for DNA analysis; RNA Extraction Protocol of LCM tissue and RT-PCR for mRNA analysis. - [Read Preparation and LCM of Paraffin Embedded Tissue Sections Protocol]

TAIL PCR Protocol - http://preuss.bsd.uchicago.edu/protocols/tail.html

Category: DNA Protocols: PCR Polymerase Chain Reaction Protocols

Mouse Caesarean Section and Fostering Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4381

Category: Mouse Protocols: Mouse Reproductive System Surgery Protocols

Protocol describes a method for Caesarean section and fostering. Caesarean section is required if the recipient of an embryo transfer or any pregnant mouse has not given birth by the delivery time normal for the particular strain. - [Read Mouse Caesarean Section and Fostering Protocol]

Isolation of High-molecular-weight DNA from Mammalian Cells Using Proteinase K and Phenol Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4036

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Isolation from Mammalian Cells Protocols

Method of choice when large amounts of mammalian DNA are required, for example, for Southern blotting (Rapid Isolation of Mammalian DNA, Rapid Isolation of Yeast DNA, Southern Blotting: Capillary Transfer of DNA to Membranes) or for construction of genomic libraries in bacteriophage {lambda} vectors. Approximately 200 µg of mammalian DNA, 100-150 kb in length, is obtained from 5 x 107 cultured aneuploid mammalian cells (e.g., HeLa cells). - [Read Isolation of High-molecular-weight DNA from Mammalian Cells Using Proteinase K and Phenol Protocol]

Method: Transfer of DNA from Bacterial Colonies to Nylon Membrane or Nitrocellulose - http://humgen.wustl.edu/hdk_lab_manual/plasmid/plsmid04.html

Category: RNA Protocols: cDNA Libraries Library

Purpose: To transfer DNA from bacterial colonies to nylon (or nitrocellulose) membrane so that a library can be screened by hybridization. Jim Howe Donis Keller Lab - [Read Method: Transfer of DNA from Bacterial Colonies to Nylon Membrane or Nitrocellulose]

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]

Mouse Embryo Transfer into Pseudopregnant Recipient Protocol - http://members.cox.net/microinjectionworkshop/technical/tnp2.html

Category: Mouse Protocols: Mouse Embryology Protocols

1-cell embryo transfer into pseudopregnant recipient female mouse. - [Read Mouse Embryo Transfer into Pseudopregnant Recipient Protocol]

Blue-Native PAGE in Plants: A Tool in Analysis of Protein-Protein Interactions - http://www.plantmethods.com/content/1/1/11

Category: Plant Biology Protocols: Plant Proteins, Peptides and Antigens

BN-PAGE has become the method of choice for the investigation of the respiratory protein complexes of the electron transfer chains of a range of organisms. It allows the separation in two dimensions of extremely hydrophobic protein sets for analysis and also provides information on their native interactions. In this review we discuss the capabilities of BN-PAGE in proteomics and the wider investigation of protein:protein interactions with a focus on its use and potential in plant science. - [Read Blue-Native PAGE in Plants: A Tool in Analysis of Protein-Protein Interactions]

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]

Nucleofection and Adoptive Transfer of Primary Mouse T lymphocytes using Transient Transfection - http://www.natureprotocols.com/2006/10/02/nucleofection_and_adoptive_tra.php

Category: Transfection Protocols: Transient Transfection Protocols

Transient transfection of genes into naïve mouse T cells using nucleofection using a modified electroporation technique. This protocol offers the possibility for rapid functional in vivo studies of targeted genes in primary mouse T cells. - [Read Nucleofection and Adoptive Transfer of Primary Mouse T lymphocytes using Transient Transfection]

PEI Nanoparticles for Targeted Gene Delivery Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/1/pdb.prot4451

Category: Gene Delivery Protocols: DNA Nanoparticles Protocols

This protocol describes the preparation of polyethylenimine (PEI)/DNA nanoparticles for targeted gene delivery. This delivery strategy improves the efficiency of gene transfer by enhancing the entry of gene vectors into the desired cells and reducing uptake by nontarget cells. We describe here methods for the conjugation of targeting peptides to PEIs, formation of DNA complexes using the conjugated PEIs or nonconjugated PEIs together with targeting peptides, and cell transfection. - [Read PEI Nanoparticles for Targeted Gene Delivery Protocol]

Oviduct Transfer Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4379

Category: Mouse Protocols: Mouse Reproductive System Surgery Protocols

Protocol describes oviduct transfer of mouse embryos. It is based on the Whittingham method, which applied a well described procedure for the rat to the mouse. It is best to practice this procedure first on a cadaver and then on an anesthetized 0.5-dpc pseudopregnant mouse using a dye solution or blue Affigel beads rather than embryos to gain experience in finding the opening of the oviduct (infundibulum). - [Read Oviduct Transfer 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]

96-Well Confirmation Yeast PCR Protocol - http://sequence-www.stanford.edu/group/yeast_deletion_project/96_well_confirmation.html

Category: Yeast Protocols: Yeast Nucleic Acid Protocols: Yeast PCR Protocols

Protocol for 96-well confirmation Yeast PCR. Includes: Clonal purification; Generate a master plate (96-well format); Making a frozen backup stock; Confirmation PCR for one Row; ORF Specific Confirmation PCR --> "A-B" primers (upstream junction); Transfer template DNA to multiwell PCR plate; Prepare and dispense master mix for A-B PCR. - [Read 96-Well Confirmation Yeast PCR Protocol]

Agrobacterium-Mediated Gene Transfer via Hypocotyls Protocol - http://wheat.pw.usda.gov/~lazo/methods/lazo/cottontr.html

Category: Plant Biology Protocols: Plant Genetics Protocols

Protocol describing Agrobacterium mediated gene transfer via hypocotyls. - [Read Agrobacterium-Mediated Gene Transfer via Hypocotyls Protocol]

Articles (1-10 of 11)

Lambda Phage Protocol Large DNA Preparation

Tubulin Polymerization Protocol using GTP and GMPCPP

Tubulin is polymerized into microtubules by incubating tubulin at 37°C with GTP. A nucleation seed is added when the purpose is to assay microtubule elongation. Tubulin can also be polymerized for the purposes of recycling the tubulin or labeling the microtubules with fluorescently labeled tubulin. Based on the protocol by Timothy Mitchison of Harvard University.

Microarray Protocol Preparation of Fluorescent DNA Probes from Human mRNA

This Microarray Protocol Preparation of Fluorescent DNA Probes from Human mRNA protocol describes the production of probes labeled with the fluorescent dyes, Cy3 and Cy5, following the synthesis of cDNA from human mRNA and the hybridization of the probes to DNA microarrays.

Probe Preparation for 22 x 40 mm DNA Microarrays

Probe Preparation for 22 x 40 mm DNA Microarrays Protocol.

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.