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Western Blot Home - http://www.molecularstation.com/protein/western-blot/

Category: Protein Protocols: Western Blot Protocol

Western Blot Home. Information on western blotting, western blot procedure and methods, western blot books, stripping protocols. - [Read Western Blot Home]

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Precision Engineering of Plant Gene Loci by Homologous Recombination Cloning in Escherichia Coli - http://www.plantmethods.com/content/1/1/6

Category: Plant Biology Protocols: Arabidopsis Genetics Protocols

Protocol for precision engineering of plant gene loci by homologous recombination cloning in Escherichia coli. Includes: Key steps in the EL250 RED-HR locus rescue and engineering procedure; Primer design and plasmid constructs; AtSTM gap-repair construct; Targeting construct backbone; Preparation of electrocompetent EL250 cells; Transformation of BAC F24o1 and induction of recombinogenic function in EL250; AtSTM locus rescue from BAC F24o1 by gap-repair HR. - [Read Precision Engineering of Plant Gene Loci by Homologous Recombination Cloning in Escherichia Coli]

Preferential Delivery of the Sleeping Beauty transposon System to Livers of Mice by Hydrodynamic i - http://www.natureprotocols.com/2007/12/06/preferential_delivery_of_the_s.php

Category: Developmental Biology: Utilizing Model Organisms Protocols

Nonviral, DNA-mediated gene transfer is an alternative to viral delivery systems for expressing new genes in cells and tissues. The Sleeping Beauty (SB) transposon system combines the advantages of viruses and naked DNA molecules for gene therapy purposes; however, efficacious delivery of DNA molecules to animal tissues can still be problematic. Here we describe the hydrodynamic delivery procedure for the SB transposon system that allows efficient delivery to the liver in the mouse. - [Read Preferential Delivery of the Sleeping Beauty transposon System to Livers of Mice by Hydrodynamic i]

Preparation and Loading of Protein Samples for Microinjection Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4656

Category: Microinjection Protocols: Microinjection of Proteins Protocols

his protocol provides methods for the preparation of protein samples and for loading them into pulled microinjection pipettes. Stock solutions of proteins are thawed, diluted (if desired), centrifuged at high speed to remove aggregates, and kept on ice until loading. Loading into micropipettes can be done using either a "front-loading" or a "backfilling" procedure. - [Read Preparation and Loading of Protein Samples for Microinjection Protocol]

Preparation and Transformation of Competent E. coli Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3942

Category: E Coli Protocols: E coli Transformation Protocols

Procedure generates competent cultures of E. coli that can be transformed at high frequencies (5 x 108 transformed colonies/µg of superhelical plasmid DNA). IMPORTANT All steps in this protocol should be carried out aseptically. - [Read Preparation and Transformation of Competent E. coli Protocol]

Preparation of Affinity-Ligand Resins by Immobilization of Dyes on Polyhydroxyl Matrices - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4208

Category: Chromatography Protocols: Dye Ligand Affinity Chromatography Protocols

Triazine dyes, such as Cibacron Blue 3GA, can be linked to a hexyl spacer arm and then immobilized on a polyhydroxyl support matrix that has been activated with either 1,1-carbonyldiimidazole or epichlorohydrin. An alternative procedure for immobilizing dyes using the direct coupling method is provided in Immobilization of Dyes on Polyhydroxyl Matrices Using the Direct Coupling Method. - [Read Preparation of Affinity-Ligand Resins by Immobilization of Dyes on Polyhydroxyl Matrices]

Preparation of Arabidopsis Chloroplasts Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4681

Category: Plant Biology Protocols: Arabidopsis Tissue Culture

It is desirable to prepare subcellular fractions, either to localize proteins or to improve the sensitivity of protein detection. This procedure describes the enrichment of chloroplasts from Arabidopsis. - [Read Preparation of Arabidopsis Chloroplasts Protocol]

Preparation of Arabidopsis Mitochondria Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4682

Category: Plant Biology Protocols: Arabidopsis Tissue Culture

It is often desirable to prepare subcellular fractions, either to localize proteins or to improve the sensitivity of protein detection. This procedure describes the enrichment of mitochondria from Arabidopsis. - [Read Preparation of Arabidopsis Mitochondria Protocol]

Preparation of B-Lymphocyte RNA for Microarray Analysis Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000009.pdf?pid=PP00000009

Category: Immunology: B Cell Protocols

Procedure permits the isolation of at least 5 µg of total RNA from a sample of purified mouse splenic B lymphocytes. The quality of the RNA is assessed by separation of an aliquot through 1% agarose and staining with ethidium bromide as described in AfCS protocol Visualization of RNA Preparations on 1% Agarose Gels. The isolated RNA is used for analysis of gene expression by microarray technology. analysis of gene expression by microarray technology. - [Read Preparation of B-Lymphocyte RNA for Microarray Analysis Protocol]

Preparation of Cells and Reagents for Flow Cytometry Protocols - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E661C3CC685613447CEC768412FDE0F&objectid=6674EA50F5C3D9A0189B376C7A89BF6B

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols

A procedure for direct and indirect staining of single-cell suspensions of lymphoid tissue or peripheral blood lymphocytes to detect cell surface membrane antigens is presented. In addition, support protocols present methods for fluorescence labeling of purified antibodies. A protocol for flow cytometric analysis of intracellular antigens in single-cell suspensions is also included. - [Read Preparation of Cells and Reagents for Flow Cytometry Protocols]

Preparation of DNA Affinity Resin - Subscription Required - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4205

Category: Chromatography Protocols

This protocol describes the preparation of concatamerized oligonucleotides and their coupling to cyanogen-bromide-activated Sepharose. The procedure uses a commercially activated resin, which can be purchased as a lyophilized powder. Keith Brocklehurst et al. - [Read Preparation of DNA Affinity Resin - Subscription Required]

Preparation of Endothelial Cells Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=E538DDD4A302A0DF2CDDD9363072C1CA&objectid=6673B460F50B90057960CE47ABB708A4

Category: Cell Biology and Cell Culture: Endothelial Cell Protocols

Endothelial cells, which line blood vessels, can be prepared from a variety of tissues. They are frequently prepared from the umbilical vein, which is relatively easy to obtain. The procedure is clearly described and provides a large population of highly purified endothelial cells. There are also methods for obtaining endothelial cells from other tissues such as fat, skin, and mucosa. These methods require special care and generate smaller populations of cells. - [Read Preparation of Endothelial Cells Protocol]

Preparation of Frozen Stocks of RAW 264.7 Cells - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000180.pdf?pid=PP00000180

Category: Cell Culture Protocols: Cell Lines Protocols

Protocol for the preparation of frozen stocks of RAW 264.7 cells. Includes: Freezing procedure and reagents. - [Read Preparation of Frozen Stocks of RAW 264.7 Cells]

Preparation of KC Nuclear Extract for In Vitro Splicing - http://www.bio.com/protocolstools/protocol.jhtml%3Bjsessionid%3D34E3CBZRBQMFZR3FQLMCFEWHUWBNSIV0?id=p521

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

Protocol for preparation of KC nuclear extract for in vitro splicing. Protocol makes 3.4 ml of extract for every 4 liter of cells (depending on initial cell concentration). Protocol includes: Procedure, Solutions, BioReagents and Chemicals and protocol hints. - [Read Preparation of KC Nuclear Extract for In Vitro Splicing]

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]

Preparing and Transforming Spheroplasts of Neurospora crassa Protocol - http://www.fgsc.net/neurosporaprotocols/How%20to%20Prepare%20Spheroplasts.pdf

Category: Fungi Protocols: Neurospora Protocols

Procedure for preparing and transforming spheroplasts of Neurospora crassa. The procedure is designed to produce a large number of spheroplasts to be stored frozen at -80°. - [Read Preparing and Transforming Spheroplasts of Neurospora crassa Protocol]

Preparing Chemically Competent Cells - http://openwetware.org/wiki/Preparing_chemically_competent_cells

Category: Microbiology: Bacterial Competent Cells Preparation E.Coli

Detailed protocol Preparation and Steps for E.Coli Competent Cells procedure. Open Net Ware. - [Read Preparing Chemically Competent Cells]

Procedure for Purifying Cell Walls from Arabidopsis Hypocotyls - http://www.plantmethods.com/content/2/1/10

Category: Plant Biology Protocols: Arabidopsis Tissue Culture

Protocol for the evaluation of cell wall preparations for proteomics: a new procedure for purifying cell walls from Arabidopsis hypocotyls. - [Read Procedure for Purifying Cell Walls from Arabidopsis Hypocotyls]

Procedure for Transfection of Mammalian Cells Using Lipofectamine - http://www.research.umbc.edu/~jwolf/m10.htm

Category: Cell Biology and Cell Culture: Cell Transfection Protocol

Lipofectamine transient tranfection of mammalian cells in IMDM cell media. Julie B. Wolf, UMBC. - [Read Procedure for Transfection of Mammalian Cells Using Lipofectamine]

Protein Coimmunoprecipitation in Arabidopsis Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4683

Category: Plant Biology Protocols: Arabidopsis Protein Peptide Protocols

Coimmunoprecipitation is one of the most useful techniques for revealing protein-protein interactions. Good negative controls to verify the specificity of the coimmunoprecipitation procedure are (1) performing the same immunoprecipitation experiment using beads coupled to the preimmune serum and (2) probing the Western blot with antibodies against protein known not to interact with the coimmunoprecipitated proteins under physiological conditions. - [Read Protein Coimmunoprecipitation in Arabidopsis Protocol]

Protocol for Conditional Ablation of stat3/socs3 Discloses the Dual Role for Reactive Astrocytes - http://www.natureprotocols.com/2006/06/30/conditional_ablation_of_stat3s.php

Category: Signalling Signal Transduction Protocols

Protocol demonstrates that reactive astrocytes play a crucial role in wound healing and functional recovery by using mice with a selective deletion of the signal transducer and activator of transcription-3 (STAT3) or suppression of cytokine signaling-3 (SOCS3) under the control of Nestin gene promoter/enhancer (STAT3Nâ€“/â€“, SOCS3Nâ€“/â€“). Procedure includes: Surgical procedures, Functional evaluation, Immunohistochemistry, In vitro migration assay. - [Read Protocol for Conditional Ablation of stat3/socs3 Discloses the Dual Role for Reactive Astrocytes]

Protocol for Real-Time RT-PCR Using SYBR Green - http://pga.mgh.harvard.edu/primerbank/PCR_protocol.html

Category: Real Time PCR

A detailed experimental procedure for real-time RT-PCR using SYBR Green I as mentioned in Wang et al 2003. - [Read Protocol for Real-Time RT-PCR Using SYBR Green]

Protocol for Trans-Well Migration Assay - http://www.natureprotocols.com/2006/08/07/transwell_migration_assay.php

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

Protocol includes procedure for Transwell in vitro migration assays. - [Read Protocol for Trans-Well Migration Assay]

Purification of Caveolae Membranes from a Plasma Membrane Fraction of Cultured Cells and Tissues - http://www.axis-shield.com/densityhome/optiprep/S19.pdf

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Membrane Isolation Protocols

The first part of the isolation procedure is a flotation through a continuous iodixanol gradient; this gradient is essentially a resolving gradient in which the caveolin-rich vesicles are concentrated in the top third of the gradient, while the predominantly caveolin-poor vesicles band in denser regions. A second discontinuous gradient is essentially a concentration gradient to band the caveolin-rich vesicles sharply at an interface. - [Read Purification of Caveolae Membranes from a Plasma Membrane Fraction of Cultured Cells and Tissues]

Purification of Nucleic Acids by Extraction with Phenol:Chloroform Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4455

Category: Nucleic Acid Purification Protocols

Protocol describes the standard method for nucleic acid purification by extraction first with phenol:chloroform (optionally containing hydroxyquiniline at 0.1%) and then with chloroform to remove any remaining phenol. The procedure takes advantage of the fact that deproteinization is more efficient when two different organic solvents are used instead of one. - [Read Purification of Nucleic Acids by Extraction with Phenol:Chloroform Protocol]

Purification of Peroxisomes and Mitochondria from Spinach Leaf by Percoll Gradient Centrifugation - http://www.plantphysiol.org/cgi/reprint/75/3/670

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Mitochondria Isolation Protocols

A procedure developed to purify simultaneously peroxisomes and mitochondria from spinach (Spirnacia olercea L.) leaf under isoosmotic and low viscosity conditions. This method involves differential centrifugation and density gradient centrifugation on four layers of Percoll. - [Read Purification of Peroxisomes and Mitochondria from Spinach Leaf by Percoll Gradient Centrifugation]

Articles (1-10 of 12)

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.

Paraffin Embedding Protocol

Paraffin Embedding Protocol for molecular profiling. This Paraffin Embedding Protocol describes the processing of the tissues into sections following ethanol fixation. Molecular profiling (MP) is a technique that is used to visualize the global patterns of RNA expression or protein expression in various cell types and disease processes.