systems Protocols

Category: Epigenetics and Methylation Protocols

The pattern and extent of DNA methylation can significantly affect the success of restriction digestions or bacterial transformations. Prokaryotic DNA may be methylated by host restriction/modification systems, while eukaryotic DNA often is methylated i - [Read A practical guide to DNA methylation Promega PDF]

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols

Protocol describes systems in which the membranes are allowed to float through a density gradient from a dense load zone. This is an ideal format. - [Read A Self-generated Gradient to Discriminate a Cytosolic or Membrane Location of Large Protein Complex]

Category: Immunology: B Cell Protocols

Describes the antigenic stimulation of in vitro antibody production by B cells and the subsequent measurement of secreted antibodies. The first protocol is a generalized system for inducing in vitro antibody production and can accommodate various types of antigens under study. Secreted antibodies can then be measured with an enzyme-linked immunosorbent assay (ELISA) or other soluble-antibody detection systems. - [Read Assays for B Lymphocyte Function Protocols]

Category: Cell Biology and Cell Culture

Cell fractionation techniques are presented for the design of gradient systems for separating one or more cell types from lavages of body cavities or from mechanically or enzymically-dissociated tissues. Includes: Preparation of cell suspension for gradient loading; Fractionation by buoyant density; Fractionation on the basis of cell size. - [Read Cell Fractionation of a Mixed Population of Cells]

Category: Microscopy Protocols: Live-Cell Imaging Protocols

Specimen chambers have had many designs published over the years describing systems that offer excellent optical properties while allowing specimens to be maintained for varying amounts of time. Ranging in complexity from the simple preparation of a sealed coverslip on a microscope slide to sophisticated perfusion chambers that enable tight control of virtually all environmental variables culture chambers are designed to to allow living specimens to be observed with minimal invasion at high res. - [Read Culture Chambers for Live-Cell Imaging]

Category: Immunology: Immunocytochemistry Protocols

Staining Tissue, Staining Cells Smeared on Slides. Positive staining controls, Negative staining controls, Immunoenzymatic Technique and FAQ. R & D Systems. - [Read Enzymatic Protocol for Immunocytochemistry]

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

Provides information on specific parameters you need to be aware of when using eukaryotic in vitro translation systems. Includes: DNA Template Considerations; Protein Labeling; Non-Radioactive Protein Labeling. - [Read Eukaryotic In Vitro Translation Systems]

Category: Immunology: Immunocytochemistry Protocols

Staining Cells in Suspension, Staining Cells Smeared on Slides. Immunofluorescent Technique. R & D Systems. - [Read Fluorescence Protocol Immunocytochemistry]

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

HUMAN EMBRYONIC STEM CELL CULTURE IN MICROFLUIDIC CHANNELS. Abhyankar et al., 2003. 7th lnternat~onal Conference on Miniaturized Chemical and Blochemlcal Analysts Systems - [Read HUMAN EMBRYONIC STEM CELL CULTURE IN MICROFLUIDIC CHANNELS]

Category: Microscopy Protocols: Electron Microscopy Protocols

In recent years, the increased sensitivity of electron detectors and the availability of low-vacuum or variable-pressure systems have allowed imaging of fresh tissue samples without the need for fixation, drying, and coating. This obviously saves a lot of time, although the image quality may not be as good as that obtained from fixed samples. However, for most applications that tend to be at a relatively low magnification, the quality can be as good as that obtained from fixed samples. - [Read Imaging of Fresh Arabidopsis Tissues in the Scanning Electron Microscope]

Category: Plant Biology Protocols

In recent years, the increased sensitivity of electron detectors and the availability of low-vacuum or variable-pressure systems have allowed imaging of fresh tissue samples without the need for fixation, drying, and coating. This obviously saves a lot of time, although the image quality may not be as good as that obtained from fixed samples. However, for most applications that tend to be at a relatively low magnification, the quality can be as good as that obtained from fixed samples. - [Read Imaging of Fresh Arabidopsis Tissues in the Scanning Electron Microscope Protocol]

Category: Microscopy Protocols: Live-Cell Imaging Protocols

Using confocal laser-scanning microscope & GFP fusion proteins in time-lapse imaging to visualize the behavior of organelles and to track membrane-bound transport intermediates that bud off from organelles. Practical issues related to construction & expression of GFP fusion proteins are discussed. Essential for optimizing the brightness and expression levels of GFP fusion proteins so that intracellular membrane-bound structures containing these fusion proteins can be readily visualized. - [Read Imaging of Organelle Membrane Systems and Membrane Traffic in Living Cells]

Category: Microscopy Protocols

The atomic force microscope (AFM) is one of the most powerful tools for determining the surface topography of native biomolecules at subnanometer resolution. The AFM can also provide insight into the binding properties of biological systems. In order to determine the specific interaction between two kinds of molecules (e.g., avidin and biotin). Includes information on principle of AFM and application of AFM. - [Read Imaging, Measuring and Manipulating Native Biomolecular Systems with the Atomic Force Microscope]

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

Useful techniques to circumvent disruption of tissue structure in the analysis of gene expression are LCM and LDM. While they require specialized microscopes and systems, they are similar in that freshly-cut frozen tissue sections can be microdissected using either a general histological stain (like H&E) or by staining with fluorescently conjugated antibodies. The LCM system by Arcturus involves... - [Read Immunofluorescent Staining for the Laser Microdissection of Individual Cells Protocol]

Category: Microscopy Protocols: Confocal Microscopy Protocols

Information on: Applications of Confocal Microscopy; Practical Instruments; Limitations of point-scanning Confocal Microscopy; Parallel beam confocal Imaging Systems. - [Read Information on Confocal Imaging]

Category: Protein Protocols: Protein Expression Protocols

KickStartâ„¢ Protein Expression Kit PDF. Dual Systems Biotech - [Read KickStartâ„¢ Protein Expression Kit PDF]

Category: Molecular Imaging: Light Microscopy

Light Microscopy - Microscopes in Cell Biology. House Ear Institute. Fluorescence microscopy, Nomarski differential interference contrast, Comparison between phase contrast and interference contrast optical systems , Interference contrast, Phase contrast, Darkfield illumination, alignment of Kohler illumination system, Protocol for using oil immersion lenses, Use of immersion oil, Calculating the final magnification on the photomicrograph, vibration, The coverslip glass, Photomicroscopy. - [Read Light Microscopy - Microscopes in Cell Biology]

Category: C. elegans Protocols

To image early cleavages and chromatin dynamics, it is convenient to use histone H2B fused to GFP or lamin::GFP. Time-lapse movies can be obtained using conventional confocal microscope systems and their included software. Early embryos dissected from transgenic hermaphrodites are placed with egg salts on agar pads. - [Read Live Imaging of Caenorhabditis elegans: Examples]

Category: Microscopy Protocols: In Vivo Imaging Protocols

Live nematodes can be studied at all levels of microscopic resolution. Data collection includes information on: Photographic Records, Video Microscopy,Multiple-Focal-Plane Time-Lapse Recording Systems and Digital Imaging. - [Read Live Imaging of Caenorhabditis elegans: Observation of Nematodes and Data Collection]

Category: RNA Protocols: mRNA Extraction Protocols

mRNA Preparation. Promega. Abstract for PolyATtract® mRNA Isolation Systems - [Read mRNA Preparation]

Category: Cytogenetics Protocols: In Situ Hybridization Protocols

Protocol for multiple-target DNA in situ hybridization with enzyme-based cytochemical detection systems. Includes: Cell preparations; Cell processing; Probe preparation; Multiple-target in situ hybridization (ISH); ISH with separate probe and target denaturation [for probes with repetitive (e.g., Alu) elements]; Post-hybridization washes; Enzyme-based cytochemical detection; etc.. - [Read Multiple-Target DNA In Situ Hybridization with Enzyme-Based Cytochemical Detection Systems Protocol]

Category: Microscopy Protocols: Optical Microscopy Protocols

Diffraction-limited optical microscopy requires that the spatial resolution of an image is limited by the wavelength of the incident light & by the numerical apertures of the condenser & objective lens systems.The development of near-field scanning optical microscopy (scanning near-field optical microscopy) has allowed for a imaging technique that retains the various contrast mechanisms afforded by optical microscopy methods while attaining spatial resolution beyond the optical diffraction limit - [Read Near-Field Scanning Optical Microscopy]

Category: Pharmacology and Toxicology Protocols: Enzyme Assays Protocols

Protocol for P450 screening system. - [Read P450 Screening Systems Protocol]

Category: Recombinant Protein Protocols: Baculovirus Recombinant Protein

PERSPECTIVES ON BACULOVIRUS EXPRESSION SYSTEMS. Advantages and disadvantages of protein expression in baculovirus . Insect Cell Culture. Frank Lab. - [Read PERSPECTIVES ON BACULOVIRUS EXPRESSION SYSTEMS]

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]