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Measurement of Green Fluorescent Protein Expression and DNA Content in Unfixed Cells Protocol - http://cyto.mednet.ucla.edu/Protocols/GFPandDNAHoechst.htm

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: Flow Cytometry Data Analysis Protocols

Protocol for measurement of green fluorescent protein expression and DNA content in unfixed cells. - [Read Measurement of Green Fluorescent Protein Expression and DNA Content in Unfixed Cells Protocol]

Measuring Protein Concentration by Western Analysis Using Enhanced Chemiluminescence Detection - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4255

Category: Western Blot Protocols: Antigen Detection Methods Protocols

Protocol describes, samples containing the target protein are deposited onto a polyvinyldifluoride (PVDF) membrane using a vacuum manifold. The immobilized protein is exposed to an antibody specific for the target protein, followed by an antibody that reacts with species-specific determinants carried by the primary antibody and is conjugated to horseradish peroxidase (HRP). - [Read Measuring Protein Concentration by Western Analysis Using Enhanced Chemiluminescence Detection]

Measuring Yeast Cell Density by Spectrophotometry Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4186

Category: Yeast Protocols

Simple method for determining the cell density of yeast cultures. - [Read Measuring Yeast Cell Density by Spectrophotometry Protocol]

Membrane Trafficking and Organelle Reagents - http://www.cshprotocols.org/cgi/content/extract/2006/29/pdb.ip23

Category: Microscopy Protocols: Live-Cell Imaging Protocols

Several common drugs, their targets, and protocols are described for studying organelle distribution and trafficking. The drugs are readily available from general suppliers, including Sigma, Roche, and Calbiochem. - [Read Membrane Trafficking and Organelle Reagents]

Metabolic Labeling of Plant Cell Cultures with K15NO3 Protocol - http://www.plantmethods.com/content/2/1/14

Category: Plant Biology Protocols: Arabidopsis Tissue Culture

Protocol for metabolic labeling of plant cell cultures with K15NO3 as a tool for quantitative analysis of proteins and metabolites. - [Read Metabolic Labeling of Plant Cell Cultures with K15NO3 Protocol]

Methylation profiles of genes utilizing newly developed CpG island methylation microarray on colorec - http://nar.oxfordjournals.org/cgi/content/full/33/5/e46

Category: DNA Microarray Protocols: DNA Methylation Microarray CpG

Kimura et al., NAR 2005. "easy-handling technology provided reproducible and precise measurement of methylated CpGs in MGMT promoter and, thus, our method may bring about a potential evolution in the handling of a variety of high-throughput DNA methylatio - [Read Methylation profiles of genes utilizing newly developed CpG island methylation microarray on colorec]

Microinjection of dsRNA into Drosophila Embryos Protocol - http://www.cshprotocols.org/cgi/content/full/2006/21/pdb.prot4516

Category: Microinjection Protocols: Microinjection of DNA Protocols

This protocol provides a description of how to introduce double-stranded RNA (dsRNA) into Drosophila embryos by microinjection. Several days of preparation are required before injections into Drosophila embryos begin. Flies must be in abundant supply for egg collection. Bombardment of embryos with dsRNA-coated gold particles (Delivery of dsRNA into Drosophila Embryos by a Gene Gun) can be used as an alternative. - [Read Microinjection of dsRNA into Drosophila Embryos Protocol]

Microinjection of dsRNA into Mouse Oocytes and Early Embryos Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/21/pdb.prot4511

Category: RNAi Technology Protocols: RNAi Mouse Embryos and Oocytes Protocols

Protocol describes how to introduce a double-stranded RNA (dsRNA) of choice into mouse oocytes or fertilized one-cell embryos by microinjection. For collection of mouse oocytes and early embryos, see Collection of Mouse Oocytes for RNAi and Collection of Early Mouse Embryos for RNAi. - [Read Microinjection of dsRNA into Mouse Oocytes and Early Embryos Protocol]

Microinjection of Mouse Zygotes Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4397

Category: Microinjection Protocols: Microinjection of DNA Protocols

This protocol describes a method for microinjection of mouse zygotes to produce transgenic mice. - [Read Microinjection of Mouse Zygotes Protocol]

Microinjection of Protein Samples Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4657

Category: Microinjection Protocols: Microinjection of Proteins Protocols

This protocol describes a method for microinjecting proteins into the nucleus or cytoplasm of adherent cells. Microinjection equipment can be obtained from a number of suppliers; this protocol has been used with the Narishige IM-200 air pressure regulator and the Leitz micromanipulator. Using this system, it is possible to microinject a constant volume within a 50% difference among cells. - [Read Microinjection of Protein Samples Protocol]

Microinjection Setup Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4396

Category: Microinjection Protocols: Microinjection Setup Protocols

This protocol describes the microinjection setup for injecting mouse zygotes to produce transgenic mice. - [Read Microinjection Setup Protocol]

Microscopic Analysis of Mitochondrial Transmembrane Potential Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4462

Category: Cell Organelle Protocols: Mitochondria Protocols

Protocol describes a method for the evaluation of mitochondrial function using the fluorochrome CMXRos. CMXRos is sequestered by actively respiring mitochondria, but washed out when the mitochondrial membrane potential is lost. This analysis can be combined with the TUNEL technique or immunocytochemistry. - [Read Microscopic Analysis of Mitochondrial Transmembrane Potential Protocol]

Mixed Oligonucleotide-primed Amplification of cDNA MOPAC Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3484

Category: PCR Protocols: cDNA Synthesis Protocols

In MOPAC, the amino-terminal and carboxy-terminal sequences of a peptide are used to design two redundant families of oligonucleotides encoding the aminoand carboxy-terminal sequences of the peptide. The primers are used either to amplify a segment of cDNA prepared by RT-PCR from a tissue known to express the protein or to amplify a segment of DNA from an established genomic or cDNA library. - [Read Mixed Oligonucleotide-primed Amplification of cDNA MOPAC Protocol]

Monosaccharide Analysis by Methanolysis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4247

Category: Chromatography Protocols: Gas Chromatography Protocols

Protocol describes monosaccharide analysis by methanolysis. The method gives more complex chromatograms than the corresponding alditol acetate method, because up to four methyl glycosides can be produced from each monosaccharide. However, the method is easier to perform. - [Read Monosaccharide Analysis by Methanolysis Protocol]

Monosaccharide Analysis of Phosphorylated Sugars by Methanolysis and Diazomethane Methylation - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4248

Category: Chromatography Protocols: Gas Chromatography Protocols

Protocol presents a method that allows rapid determination of the monosaccharide composition of glycans, glycoproteins, and proteoglycans that contain (or are suspected to contain) phosphorylated sugars. (For samples that do not contain phosphorylated sugars see Monosaccharide Analysis by Methanolysis.) - [Read Monosaccharide Analysis of Phosphorylated Sugars by Methanolysis and Diazomethane Methylation]

Monosaccharide Composition Analysis: Alditol Acetates Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4246

Category: Chromatography Protocols: Gas Chromatography Protocols

Protocol describes how to determine the monosaccharide composition of glycans, glycoproteins, or proteoglycans by hydrolyzing the sample to monosaccharides and converting them to alditols, then performing acetylation to make them volatile compounds and analysis by gas chromatography (GC) or gas chromatography coupled with mass spectrometry (GC-MS). - [Read Monosaccharide Composition Analysis: Alditol Acetates Protocol]

Mounting Samples in DPX Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4460

Category: Cell Culture Protocols: Cell Fixing Protocols

Mounting media for immunohistology must be compatible with the detection method used. A suitable non-aqueous mounting medium is DPX. - [Read Mounting Samples in DPX Protocol]

Mounting Samples in Gelvatol or Mowiol Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4461

Category: Cell Culture Protocols: Cell Fixing Protocols

Mounting media for immunohistology must be compatible with the detection method used. Suitable aqueous media are made from Gelvatol or Mowiol. - [Read Mounting Samples in Gelvatol or Mowiol Protocol]

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]

Mouse Castration Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4387

Category: Mouse Protocols: Mouse Reproductive System Surgery Protocols

Protocol describes castration of male mice, which is used to eliminate testicular hormones and/or to obtain testes for analysis without sacrificing the male. Castration is performed in a similar manner to vasectomy. - [Read Mouse Castration Protocol]

Mouse Nephrectomy Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4386

Category: Mouse Protocols: Mouse Organ Removal Surgery Protocols

Protocol describes surgical removal of a mouse kidney to provide tissue for analysis. - [Read Mouse Nephrectomy Protocol]

Mouse Splenectomy Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4385

Category: Mouse Protocols: Mouse Organ Removal Surgery Protocols

Protocol describes removal of a mouse spleen to provide tissue for analysis. - [Read Mouse Splenectomy Protocol]

Multiplex Real-Time PCR Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4113

Category: PCR Protocols: Real-Time PCR Protocols

In multiplex real-time PCR, different sets of primers with different labels are used to amplify separate genes from the template DNA in one tube. This protocol uses LUX (Light Upon eXtension) primers from invitrogen. FAM (6-carboxy-fluorescein) is used to label the gene of interest, and JOE (6-carboxy-4', 5'-dichloro-2',7'-dimethoxy-fluorescein) is used to label a housekeeping gene as an internal control to normalize between different reactions. - [Read Multiplex Real-Time PCR Protocol]

Mutagenic PCR Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4143

Category: Cloning Protocols: Mutagenesis Protocols

Protocol for mutagenic PCR. - [Read Mutagenic PCR Protocol]

Mutation Detection by Solid-Phase Chemical Cleavage of Mismatches Using Radioactive Probes Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/14/pdb.prot4120

Category: Genetics and Genomics: Genotyping Protocols: Mutation Detection Protocols

This protocol is the first of two describing solid-phase chemical cleavage of mismatch (SpCCM), a method for the detection of mutations in genomic DNA. DNA fragments up to 2 kb in length can be analyzed to determine the position (within 10-15 bp) and identity of the mismatched nucleotide. - [Read Mutation Detection by Solid-Phase Chemical Cleavage of Mismatches Using Radioactive Probes Protocol]

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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.

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