analysis Protocols

Category: Primary Cell Isolation and Culture Protocols: Laser Capture Microdissection Protocols: Tissue Preparation for LCM Protocols

LCM isolates specific cells or tissues from samples mounted on microscope slides. The samples are viewed through a thermoplastic film that is attached to a microcentrifuge tube lid. Localized heat, caused by the application of a laser pulse, fuses the membrane to the cells of interest, which can then be harvested for further analysis. RNA and proteins can be purified from the isolated cells, allowing detailed analysis of gene expression. This protocol is divided into three stages. - [Read (LCM): Preparation and Sectioning of Frozen Tissue Blocks and Purification of RNA from Isolated Cel]

Category: DNA Microarray Protocols

Hegde et al., Array Fabrication, microarray Printing, Probe Preparation and Hybridization, RNA Extraction, RNA labeling, Data Collection, Normalization, and Analysis.Institute for Genomic Research, Rockville, MD - [Read A Concise Guide to cArray Hybridization Procedures - DNA Microarray Analysis]

Category: DNA Microarray Protocols

Micro Array Fabrication, Probe Preparation and Hybridization, and Data Collection, Normalization and Analysis. Hegde, et al. biotechniques 2000. - [Read A Concise Guide to cDNA Microarray Analysis – II PDF.]

Category: Genetics and Genomics: Cancer Genetics Protocols: Loss of Heterozygosity Protocols

DNA for analysis is purified using salt precipitation. The method is gentle, limits the breakage of the long chromosomal strands, and avoids the use of phenol and chloroform. It is suitable for use with cultured cells, breast tumor tissue that has been subjected to hormone receptor analysis, and blood samples. The loss of heterozygosity assay is performed using a multiplex PCR, in which one of each primer pair is labeled with a different fluorophor. - [Read A Multiplex PCR Method to Define a Narrow Deleted Chromosomal Region of a Tumor Genome]

Category: DNA Protocols: EMSA DNA-Protein Protocols

A non-radioactive electrophoretic mobility shift assay based on the digoxigenin detection system had been developed and applied to the analysis of the interaction between the heat shock transcription factor and the heat shock element of N. crassa. (Bioch - [Read A non-radioactive electrophoretic mobility shift assay for the detection of heat shock element (HSE)]

Category: Plant Biology Protocols: Plant Genetics Protocols

For analysis of metaphase chromosomes, any tissue containing dividing cells can be used: Root tips from young seedlings, from newly grown roots at the edge of plant pots or hydroponic culture are all suitable. Alternatively, flower buds, anthers, carpels or leaf or apical meristems can be used. Includes metaphase arresting reagents. - [Read Accumulation and Fixation of Plant Metaphase Chromosomes Protocol]

Category: Microarray Protocols: Microarray Chip Protocols

Protocol for agilent yeast ChIP-on-chip analysis. - [Read Agilent Yeast ChIP-on-chip Analysis Protocol]

Category: DNA Microarray Protocols: DNA Microarray Software

DNA Microarray Software. Data Analysis, Another Microarray Database AMAD, Cluster, Gal File Maker, ScanAlyze. DeRisi Lab. - [Read AMAD: Another Microarray Database.]

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: DNA Staining Protocols for Flow Cytometry

Simple and universally applicable methods for staining fixed cells are presented, as are methods that utilize detergents and proteolytic treatment to permeabilize cells. Additionally, supravital cell staining with Hoechst 33342, which is primarily used for sorting live cells for subsequent culturing based on DNA-content differences, is also described. Also presented are methods for staining of cell nuclei isolated from paraffin-embedded tissues, and deconvolution of DNA-content-frequency... - [Read Analysis of Cellular DNA Content by Flow Cytometry Protocol]

Category: DNA Protocols: Genomic DNA Library Protocols

Amplification and sequencing of the exon-trapped products. - [Read Analysis of Clones for Exon Trapping and Amplification]

Category: Molecular Biology Protocols: Restriction Enzyme Digestion

Very detailed class protocol for Analysis of DNA by Restriction Enzyme Cleavage and Agarose Gel Electrophoresis. David Ng, UBC. - [Read Analysis of DNA by Restriction Enzyme Cleavage and Agarose Gel Electrophoresis]

Category: Immunohistochemistry Protocols: Fixation Protocols

Protocol for analysis of DNA content and surface immunophenotype using gentle ethanol fixation techniques. - [Read Analysis of DNA Content and Surface Immunophenotype Protocol Using Ethanol Fixation]

Category: Cell Biology and Cell Culture: Apoptosis Protocols: Apoptosis Analysis

Analysis of DNA Fragmentation Using Agarose Gel Electrophoresis Shailaja Kasibhatla et al. This protocol provides a qualitative method for assessing cell death by detecting DNA fragments using agarose gel electrophoresis. One of the classic features of apoptosis is the cleavage of the genomic DNA into oligonucleosomal fragments represented by multiples of 180-200 bp. Visualizing these fragments can aid in characterizing an apoptotic event. May be combined with more quantitative methods. - [Read Analysis of DNA Fragmentation Using Agarose Gel Electrophoresis (Subscription Required)]

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: DNA Staining Protocols for Flow Cytometry

This protocol provides a method for analysis of DNA fragmentation using flow cytometry after propidium iodide (PI) labeling of apoptotic nuclei. Apoptotic nuclei stain less than their normal counterparts because of diffusion of low-molecular-weight fragments out of the cells and/or chromatin condensation. - [Read Analysis of DNA Fragmentation Using Propidium Iodide (PI) Fluorescence of Individual Nuclei Protocol]

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: DNA Staining Protocols for Flow Cytometry

This protocol provides a method for analysis of DNA fragmentation using flow cytometry after propidium iodide (PI) labeling of fixed apoptotic cells. Flow cytometry reveals apoptotic nuclei in the subdiploid region of the cell cycle histogram... - [Read Analysis of DNA Fragmentation Using Propidium Iodide (PI) Staining After Ethanol Fixation Protocol]

Category: Cell Biology and Cell Culture: Apoptosis Protocols: Apoptosis Analysis

Analysis of DNA Fragmentation Using the JAM Assay. By Shailaja Kasibhatla et al., The JAM assay is based on labeling nuclear DNA of cycling cells with [3H]thymidine and harvesting samples on glass fiber filters. Apoptosis will generate DNA fragments small enough to pass through the glass fiber filter, resulting in decreased radioactivity of the particular sample. Cell-mediated cytotoxicity or cell killing mediated by cytotoxic T lymphocytes (CTL) can also be measured by this technique. - [Read Analysis Of DNA Fragmentation Using The JAM Assay (Subscription Required)]

Category: PCR Protocols: Bisulfite Conversion Based PCR Protocols

Protocol for the analysis of DNA methylation using bisulphite sequencing. Method allows precise analysis of methylation in a certain region by converting all nonmethylated cytosines into tymines, while methylated cytosines remain unchanged.  This method requires small amount of genomic DNA and therefore seems to be very useful for the analysis of clinical samples, where the material amount is limited.
- [Read Analysis of DNA Methylation using Bisulphite Sequencing Protocol]

Category: PCR Protocols: Bisulfite Conversion Based PCR Protocols

Protocol for analysis of DNA methylation using SnuPE. - [Read Analysis of DNA Methylation using SnuPE Protocol]

Category: Cell Biology and Cell Culture: ES Stem Cell Biology and Culture

Analysis of ES Cell Clones by Mini-Southern. Bradley's Lab, Texas. - [Read Analysis of ES Cell Clones by Mini-Southern]

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

Provides several approaches to flow cytometry data analysis. Frequency determinations based on analysis of single-parameter fluorescence histograms and dual-parameter contour plots are presented. Steps are described for calculating values for signal-to-noise ratios when logarithmic amplification is used for data collection. - [Read Analysis of Flow Cytometry Data Protocol]

Category: Protein Protocols: Protein Trafficking

Protocol is based on methods for the resolution of GLUT4 containing vesicles and the identification of phosphoinositide kinase containing vesicles in 3T3-L1 adipocytes. They may have a wider application to any low-medium density membranes. Protocol incorporates the strategy of using a low density microsome fraction as the gradient input, commonly used in GLUT 4 studies that may have a wider application to other investigations. - [Read Analysis of Membrane Trafficking and Intracellular Signaling in Self-Generated Iodixanol Gradients]

Category: Epigenetics and Methylation Protocols: Bisulfite Treatment Methylation Assay Protocol

This method allows precise analysis of methylation in a certain region by converting all nonmethylated cytosines into tymines, while methylated cytosines remain unchanged. Dr. A. Gratchev Methods.info - [Read Analysis of methylation using bisulphite sequencing]

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]

Category: Molecular Biology Protocols: Carbohydrate Protocols

High performance liquid affinity chromatography (HPLAC) is a useful procedure to investigate he interactions between carbohydrate binding protein and their ligands. Technical requirements are similar to conventional HPLC. HPLAC can screen and separate natural ligands from complex biological mixtures. WeiTong Wang~GlycoTech Corporation, Rockville, Maryland - [Read Analysis of Oligosaccharide Ligands by High Performance Liquid Affinity Chromatography]

Category: HPLC Protocols

Analysis of Oligosaccharide Ligands by High Performance Liquid Affinity Chromatography Protocol. Glycotech. - [Read Analysis of Oligosaccharide Ligands by High Performance Liquid Affinity Chromatography]