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Antibiotic and Selection Reagent Usage for Resistant E. coli Cells PDF - http://www.atcc.org/pdf/AntibioticTable.pdf

Category: Microbiology: Antibiotics Concentration in Media LB

Antibiotics, ampicillin, choramphenicol, G418, X-Gal and IPTG also stock concentrations. ATCC. - [Read Antibiotic and Selection Reagent Usage for Resistant E. coli Cells PDF]

Antisense Oligonucleotide Transfection of RAW 264.7 Cells with FuGENE 6 in a 24-Well Dish - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000261.pdf?pid=PP00000261

Category: Transfection Protocols: Stable Transfection Protocols

The AfCS is utilizing antisense technology to manipulate signaling protein expression in the RAW 264.7 macrophage-like cell line. This can be achieved by the transfection of gene-specific antisense oligonucleotides (ASOs). The following procedure involves the transfection of ASOs into RAW 264.7 cells using FuGENE 6 transfection reagent. Subsequently, the isolated total RNA or protein from these transfected cells can be used to assess the level of mRNA or protein knockdown, respectively. - [Read Antisense Oligonucleotide Transfection of RAW 264.7 Cells with FuGENE 6 in a 24-Well Dish]

Binding Antibodies to Tissue Sections Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4331

Category: Antibody Protocols: Antibody Labeling Protocols

Once tissues are fixed and permeabilized, the antibodies are added. These antibodies can be labeled directly or detected by a labeled secondary reagent. For indirect detection, any reagent that binds specifically to the primary antibody can be "tagged" and used to locate the antibody. The possible reagents include anti-immunoglobulin antibodies, protein A or G, or, if the first antibody is labeled with biotin, streptavidin. They can be labeled with enzymes or gold. - [Read Binding Antibodies to Tissue Sections Protocol]

Calcium Flux Analysis Protocol for Flow Cytometry - http://www.niehs.nih.gov/research/atniehs/core/fc/protocols/calcium.cfm

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols

Protocol for calcium flux analysis. Includes: Reagent Preparation; Sample Preparation; Cell Surface Staining; Instrument Set up; Optimizing the Instrument Settings; Recording Data to Apply Compensation; Recording Experimental Data. - [Read Calcium Flux Analysis Protocol for Flow Cytometry]

Cell Viability Assays that Measure Metabolic Capacity Protocol - http://www.promega.com/paguide/chap4.htm#title3

Category: Cell Biology and Cell Culture: Cell Viability Protocols

The CellTiter-Blue® Cell Viability Assay uses an optimized reagent containing resazurin. The homogeneous procedure involves adding the reagent directly to cells in culture at a recommended ratio of 20µl of reagent to 100µl of culture medium. - [Read Cell Viability Assays that Measure Metabolic Capacity Protocol]

Curing Strains of Endogenous 2-Micron Plasmid Protocol - http://ygac.med.yale.edu/mtn/reagent/avail_reagents/curing2micron_info.stm

Category: Yeast Protocols: Yeast Genetics Protocols

Protocol for curing strains of endogenous 2-micron plasmid. - [Read Curing Strains of Endogenous 2-Micron Plasmid Protocol]

Cytokine ELISPOT Protocol - http://www.ebioscience.com/ebioscience/appls/ELISPOT.htm#protocol

Category: ELISA Protocols: ELISpot Protocols

Protocol for cytokine ELISPOT. Includes: Description; Stability; Contents of the ELISPOT Ready-SET-Go! Reagent Set; ELISPOT Method; Cytokine ELISPOT Buffers. - [Read Cytokine ELISPOT Protocol]

Detection of Recombinant Proteins by ELISA - http://omrf.ouhsc.edu/~frank/ELISA.html

Category: Recombinant Protein Protocols

ELISA detection of preparation of bacterially expressed protein. ELISA REAGENT RECIPES. Frank Lab. - [Read Detection of Recombinant Proteins by ELISA]

Determining Number of Live and Dead Cells in Cell Population: Cytotoxicity Assay Protocol - http://www.promega.com/paguide/chap4.htm#title4

Category: Cell Biology and Cell Culture: Cell Viability Protocols

Cell-based assays are important tools for contemporary biology and drug discovery because of their predictive potential for in vivo applications.This assay gives ratiometric, inversely proportional values of viability and cytotoxicity (Figure 4.15) that are useful for normalizing data to cell number. Also, this reagent is compatible with additional fluorescent and luminescent chemistries. - [Read Determining Number of Live and Dead Cells in Cell Population: Cytotoxicity Assay Protocol]

Dual Luciferase Reporter System Protocol (PROMEGA) - http://www.biochem.northwestern.edu/ibis/morimoto/research/Protocols/II.%20Eukaryotes/F.%20Luciferase%20Assays/2b.Dual%20luciferase.pdf

Category: Reporter Gene Assays: Luciferase Assay Protocols

Protocol for dual luciferase reporter system. Includes: Transfections; Cell Lysis; Reagent Preparation; Luminescence measurement; Wash injectors. - [Read Dual Luciferase Reporter System Protocol (PROMEGA)]

Extraction and Purification of Total RNA using TRIzol OR TRI Reagent Protocol - http://www.animal.ufl.edu/hansen/Protocols/RNA_extraction.htm

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols

Protocol for extraction and purification of total RNA using TRIzol OR TRI reagent. Includes: Homogenization for Cell Suspensions; Phase Separation; RNA Precipitation; RNA Wash; Redissolving the RNA; Determination of RNA Concentration and Purity; Preparation of Rnase-free water. - [Read Extraction and Purification of Total RNA using TRIzol OR TRI Reagent Protocol]

Immunoaffinity Purification: Coupling Antibodies to Protein A or G Bead Columns Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4303

Category: Chromatography Protocols: Immunoaffinity Chromatography Protocols

Protocol describes how antibodies are bound to protein A or protein G beads and are then crosslinked to the matrix via a bifunctional coupling reagent called dimethyl pimelimidate (DMP). - [Read Immunoaffinity Purification: Coupling Antibodies to Protein A or G Bead Columns Protocol]

Lowry Protein Assay - http://www.animal.ufl.edu/hansen/protocols/LOWRY.htm

Category: Protein Protocols: Lowry Protein Assay

Lowry Protein Assay. The Lowry procedure is one of the most venerable and widely-used protein assays, being first described in 1951 [Lowry et al., J. Biol. Chem. 193: 265-275 (1951)]. Under alkaline conditions, copper complexes with protein. When folin phenol reagent (phospho-molybdic-phosphotungstic reagent) is added, the Folin-phenol reagent binds to the protein. Bound reagent is slowly reduced and changes color from yellow to blue. P.J. Hansen, Dept. of Animal Sciences, University of Florida. - [Read Lowry Protein Assay]

Luciferase Assay 24 Well Plate (Promega System) - http://www.biochem.northwestern.edu/ibis/morimoto/research/Protocols/II.%20Eukaryotes/F.%20Luciferase%20Assays/2a.single%20luciferase.pdf

Category: Reporter Gene Assays: Luciferase Assay Protocols

Luciferase assay using a 24 well plate. Includes: Cell Lysis; Reagent Preparation; Luminescence measurement; Wash injector; Protein quantitation; Data Analysis. - [Read Luciferase Assay 24 Well Plate (Promega System)]

Lysis and Protein Extraction from WEHI-231 Cells with TriPure Isolation Reagent Protocol - http://www.signaling-gateway.org/data/cgi-bin/ProtocolFile.cgi/afcs_PP00000162.pdf?pid=PP00000162

Category: Chromatography Protocols: Immobilized Metal Ion Affinity Chromatography Protocols

Protocol is the first step in a three-step process for the preparation and enrichment of phosphopeptides using immobilized metal affinity chromatography (IMAC) for the identification of the phosphopeptides by LC-MS/MS. This procedure is used to prepare protein extracts from WEHI-231 cells. This preparation method provides total cellular protein samples that are free of contaminating nucleic acids. - [Read Lysis and Protein Extraction from WEHI-231 Cells with TriPure Isolation Reagent Protocol]

MTT Cell Proliferation Assay Protocol - http://www.atcc.org/pdf/30-1010k.pdf

Category: Cell Biology and Cell Culture: Cell Viability Protocols

The MTT Cell Proliferation Assay measures the cell proliferation rate and conversely, when metabolic events lead to apoptosis or necrosis, the reduction in cell viability. The number of assay steps has been minimized as much as possible to expedite sample processing. The MTT Reagent yields low background absorbance values in the absence of cells. Includes: Determining optimal cell counts, performing an assay, data interpretations and troubleshooting. - [Read MTT Cell Proliferation Assay Protocol]

Mycoplasma Elimination Reagent Protocol - http://www.biovalley.fr/anglais/biology/mynang.htm

Category: Cell Culture Protocols: Cell Culture Contamination Prevention and Removal Protocols

For both biological and economical reasons, it is important to eliminate mycoplasmas from cell cultures being used for basic research, diagnosis, and biotechnological production. The most commonly used method for elimination, inactivation, or suppression of mycoplasmas in cell cultures is treatment with antibiotics. In general, antibiotic therapies do not result in long-lasting, successful elimination. Also, the cytotoxic properties of antibiotics can cause undesirable side effects on cells. - [Read Mycoplasma Elimination Reagent Protocol]

Preparation of RNA from Paraffin-Embedded Fixed Tissue Using Trizol Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4103

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols: RNA Isolation fromTissues or Cultured Cells Protocols

The method uses Trizol, a phenol-base reagent produced by Invitrogen. - [Read Preparation of RNA from Paraffin-Embedded Fixed Tissue Using Trizol Protocol]

Protocol for Luciferase Assay for In Vitro Detection - http://aesop.rutgers.edu/~lamlab/lucassayprotocol.html

Category: Reporter Gene Assays: Luciferase Assay Protocols

Protocol for luciferase assay for in vitro detection. Protocol includes: Before cell lysate preparation; Cell lysate preparation; Protocol for manual luminometers; Protocol for plate reading luminometer; Preparation of protein assay reagent; Protein standards. - [Read Protocol for Luciferase Assay for In Vitro Detection]

Purification of RNA from Animal Cells Using Trizol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4104

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols: RNA Isolation fromTissues or Cultured Cells Protocols

Protocol for purification of RNA from animal cells using Trizol. Trizol is a phenol-based reagent produced by Invitrogen. - [Read Purification of RNA from Animal Cells Using Trizol]

Purification of RNA from Plant Tissue Using the Concert Plant Reagent - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4106

Category: Plant Biology Protocols: Plant DNA RNA Isolation Protocols

This protocol is not phenol-based, but does require the addition of chloroform. The Concert Plant Reagent is intended for the isolation of RNA from a wide variety of plant tissues including blue spruce needles, potato tuber etc. - [Read Purification of RNA from Plant Tissue Using the Concert Plant Reagent]

Quantitative GUS Activity Assay in Intact Plant Tissue Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/3/pdb.prot4688

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

The activity of ß-glucuronidase (GUS) can be accurately determined in intact plant tissue using 4-methylumbelliferyl ß-D-glucuronide (4-MUG) as a substrate. Upon hydrolysis by GUS, the fluorochrome 4-methyl umbelliferone (4-MU) is produced. This method is based on the permeability of both 4-MUG and 4-MU through plant tissue. It consists of incubation of the tissue with the reagent and quantification of the fluorescence emitted by 4-MU in the solution. GUS activity in each sample can be... - [Read Quantitative GUS Activity Assay in Intact Plant Tissue Protocol]

Real-Time RT-PCR: cDNA Synthesis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4114

Category: PCR Protocols: Real-Time PCR Protocols

Protocol uses the Superscript II First-Strand Synthesis system for the generation of cDNA from total RNA. RNA purified using TRIzol reagent (Invitrogen) or the methods described in Preparation of RNA Using Guanidinium Isothiocyanate/Cesium Chloride Ultracentrifugation, Preparation of RNA from Paraffin-Embedded Fixed Tissue. - [Read Real-Time RT-PCR: cDNA Synthesis Protocol]

Real-Time RT-PCR: cDNA Synthesis Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4114

Category: PCR Protocols: Reverse Transcriptase RT-PCR Protocols

SNP Genotyping System - http://www.promega.com/tbs/tm053/tm053.pdf

Category: Genetics and Genomics: Genotyping Protocols

SNP Genotyping System. Includes: Protocol for Manual READIT® Assay in Multiwell Plates; Preparation of L/L Reagent; Preparation of Heating Block; CIAP/Exo Treatment; Preparation of Interrogation Probe Solutions; Template Denaturation; Sample Interrogation; Protocol for Manual Plate Reading Luminometers; Protocol for Injecting Plate Reading Luminometers; Protocol for Automated READIT® Assay with the Biomek® 2000 Instrument. - [Read SNP Genotyping System]

Articles (1-7 of 7)

Single Stranded Plasmid DNA Isolation Protocol

A Single Stranded Plasmid DNA Isolation Protocol describing the production and isolation of single-stranded DNA (ssDNA) using bacteriophagemid-containing bacteria and helper phage. Infection of the host cells with helper phage allows for packaging of ssDNA into bacteriophage. The ssDNA can then be isolated from phage particles.

Acid Washing Microinjection Pipettes Protocol

Acid Washing of Glass microinjection pipettes protocol.

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.

3' Rapid Amplification of cDNA Ends RACE Using PCR Protocol

3' Rapid Amplification of cDNA Ends RACE Using PCR Protocol. This protocol contains the steps for 3' end rapid amplification of mRNA by PCR. The first-strand cDNA is synthesized from total or poly(A+) RNA by priming from the poly-A tail of the mRNA using a oligo (dT) adaptor primer. The cDNA is then amplified via PCR using a gene-specific primer and an adaptor primer.

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.

Picking Transfected ES Cell Clones Protocol

This protocol a protocol on how to generate transfected embryonic stem (ES) cell clones. The previous protocol in this series is the Protocol for Electroporation of ES cells. The next protocol in the series is the Protocol on Disaggregation, Expansion, and Freezing of Transfected ES Clones.

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.

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