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Analysis of Mitochondrial Membrane Potential with the Sensitive Fluorescent Probe JC-1 - http://www.cyto.purdue.edu/flowcyt/research/cytotech/apopto/data/cossar1/cossariz.htm

Category: Cell Organelle Protocols: Mitochondria Protocols

The technique of JC-1 staining has been developed with the intent to detect DY in intact, viable cells. For this purpose JC-1 acts as a marker of mitochondrial activity, since the formation of J-aggregates, which give red emission, is reversible. Cells with high DY are those forming J-aggregates, thus showing high red fluorescence. On the other hand, cells with low DY are those in which JC-1 maintains (or re-acquire) monomeric form, thus showing only green fluorescence. - [Read Analysis of Mitochondrial Membrane Potential with the Sensitive Fluorescent Probe JC-1]

Antibody Addition to Drosophila Specimens and Detection Using Enzyme-Linked Reagents Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/23/pdb.prot4526

Category: Drosophila Protocols

Enzyme-linked reagents give excellent sensitivity and use a simple light microscope for detection. A range of enzymes is available, but for staining in situ, horseradish peroxidase will suit most needs. Diaminobenzidine (DAB) is one of the most sensitive substrates for horseradish peroxidase. It yields an intense brown product that is insoluble in both water and alcohol. It can be made more sensitive by adding metal salts such as cobalt or nickel to the substrate solution. - [Read Antibody Addition to Drosophila Specimens and Detection Using Enzyme-Linked Reagents Protocol]

Bioassay for Determining Itraconazole Levels in Blood - http://www.aspergillus.org.uk/indexhome.htm?secure/laboratory_protocols/./itrabio.html~main

Category: Fungi Protocols

Serum concentrations of itraconazole should be measured in patients receiving this drug to ensure that therapeutic concentrations are being achieved. This is necessary as drug absorption can be variable, and levels may be lowered by interactions with other drugs. The assay will give an indication of whether suitable blood levels have been achieved. - [Read Bioassay for Determining Itraconazole Levels in Blood]

DNA IQ Isolation of Genomic DNA from Stains and Buccal Swabs Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4098

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: Genomic DNA Isolation Protocols

In this protocol, the DNA-binding capacity of Wizard MagneSil particles is used to capture and release a consistent amount of DNA (100 ng) across a wide range of samples. At the end of the procedure, the DNA is eluted into 100 µl Elution Buffer to give a final concentration of 1 ng/µl, relieving the need for postpurification DNA quantitation. - [Read DNA IQ Isolation of Genomic DNA from Stains and Buccal Swabs Protocol]

Laser-capture Microdissection Protocol - http://www.nature.com/nprot/journal/v1/n2/full/nprot.2006.85.html#a1

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

LCM technology can harvest the cells of interest directly or can isolate specific cells by cutting away unwanted cells to give histologically pure enriched cell populations. A variety of downstream applications exist: DNA genotyping and loss-of-heterozygosity (LOH) analysis, etc. Protocol provides a thorough description of LCM techniques, with an emphasis on tips and troubleshooting advice derived from LCM users. The total time required to carry out this protocol is typically 1–1.5 h. - [Read Laser-capture Microdissection Protocol]

Maxam Gilbert Sequencing for DnaseI Footprinting Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/M2.html

Category: DNA Protein Interactions Protocols: Dnase Footprinting Assay Protocols

Quick protocol to give a representative G+A ladder for DnaseI footprint experiments. - [Read Maxam Gilbert Sequencing for DnaseI Footprinting Protocol]

Presenting Exogenous Antigen to T Cells Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E6631B80A848F54654FC872A48DB060&objectid=6675843EDB95D0444CF7AA285D1714C8

Category: Immunology: Mononuclear Cell Isolation Protocols: T-Cell Isolation Protocols

Protocols utilize T hybridomas to detect expression of peptide-MHC complexes, since these cells provide the most convenient, consistent, and flexible T cell readout systems for these purposes. If desired, antigen-specific T cell clones can be used in lieu of T hybridoma cells, but T cell clones often give poorer responses than T hybridomas to fixed APCs due to fixation-induced loss of costimulator function. - [Read Presenting Exogenous Antigen to T Cells Protocol]

Protocol: Bioassay for Determining Voriconazole Levels in Blood - http://www.aspergillus.org.uk/indexhome.htm?secure/laboratory_protocols/./BIOASSAY.html~main

Category: Fungi Protocols

Serum concentrations of voriconazole should be measured in patients receiving this drug to ensure that therapeutic levels are being achieved. The assay will give an indication of whether suitable blood levels have been achieved. - [Read Protocol: Bioassay for Determining Voriconazole Levels in Blood]

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]

Serial Analysis Of Gene Expression (SAGE) - http://www.embl-heidelberg.de/info/sage/

Category: Genetics and Genomics: Cancer Genetics Protocols

SAGE is a new method that has been invented at Johns Hopkins University in USA to give scientists an overview of a cell’s complete gene activity. It works by capturing RNAs, identifying them and counting them. By comparing different types of cells, the researchers hope to generate profiles that will help them understand healthy cells and what goes wrong during diseases. Includes: How SAGE works and Steps of SAGE. - [Read Serial Analysis Of Gene Expression (SAGE)]

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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.

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 Ligation Protocol

The DNA Ligation protocol described here contains the steps required to join together using ligase enzyme both plasmid DNA and insert DNA fragments in order to create a new plasmid. This new ligated plasmid can be transformed after into competent bacteria to produce DNA for mini, midi or maxi-prep isolation.

Electrotransformation of BMH 81-17mut S for Isolating Site-Directed dsDNA Mutants

This protocol describes the electroporation of the BMH 81-17 mut S strain that is recommended for tranformation of the site directed mutagenesis of dsDNA (See Protocol on Site-Directed Mutagenesis on Double Stranded DNA). BMH 81-17 mut S are a mismatch repair defective (mut S) Escherichia coli strain. The probability that the two mutations will cosegregate during the first round of DNA replication is increased in this strain.

Vector Design Protocol for Transgenic Mice Generation

The protocol gives general considerations for the design of targeting vectors for transgenic mice. The protocol shares tips in the design of knock-out and knock-in vectors and some of their strategies for producing homologously recombined embryonic stem cells.

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