alternative Protocols

Category: Pharmacology and Toxicology Protocols: Cytotoxicity Protocols: Cytotoxicity Assays in Cell Culture Protocols

Protocol descibes the use of L929 mouse fibroblast cells cultured in vitro in an agarose overlay assay to assess the toxicity of test substances.  The assay may be useful in assessing the irritation potential of test substances (e.g. surfactant-based products) as an alternative to the Draize rabbit eye test. - [Read Agarose Overlay Assay Protocol]

Category: Yeast Cell Culture Protocols: Yeast Sporulation Protocols

Protocol for an alternative method for sporulation for BY4743. - [Read Alternate Sporulation Method for BY4743 Protocol]

Category: Cell Biology and Cell Culture: Cell Culture Growth Media: Yeast Cell Culture Growth Media Protocols

Protocol for bird Medium: an alternative to Vogel Medium. - [Read Bird Medium Protocol: An Alternative to Vogel Medium]

Category: Cell Biology and Cell Culture

The protocol presented in this Application Sheet uses an alternative strategy to sedimentation on to a density barrier, that is to adjust the density of whole blood to a value just greater than the cells of interest and allow them to float to the surface. - [Read C8 Isolation of bovine peripheral blood mononuclear cells by flotation.]

Category: Cytogenetics Protocols

This CGH Protocol is used for DNA of good quality when available in sufficient amounts. We usually do replicate hybridizations using samples labeled "inversely" (reversing the label for test and normal DNA's). If appreciable artifact occurs, then alternative labels are tried. - [Read CGH of Direct Labeled Test DNA vs Normal DNA Protocol]

Category: Cytogenetics Protocols

Protocol for color changing karytoping (CCK): an M-FISH/SKY alternative. Includes: Slide preparation and hybridization; Probes, dyes and antibodies; Probe detection and image capture; Image processing. - [Read Color Changing Karyotyping (CCK) Protocol: An M-FISH/SKY Alternative]

Category: Nucleic Acid Purification Protocols

Ultrafiltration is an alternative to ethanol precipitation for the concentration and desalting of nucleic acid solutions.  It requires no phase change and is particularly useful for dealing with very low concentrations of nucleic acids.  This protocol describes the use of the Microcon cartridge, a centrifugal ultrafiltration device, to concentrate and desalt nucleic acid solutions. - [Read Concentrating and Desalting Nucleic Acids with Microconcentrators Protocol]

Category: RNAi Technology Protocols: RNAi Mammalian Cells Protocols

Protocol describes the use of western blotting to analyze the RNAi-induced depletion of target protein from mammalian cells.  Immunofluorescence can be used as an alternative. - [Read Detection of RNAi-Induced Protein Knockdown in Mammalian Cells by Western Blotting Protocol]

Category: Western Blot Protocols

Protocol describes the use of western blotting to analyze the RNAi-induced depletion of target protein from mammalian cells.  Immunofluorescence can be used as an alternative. - [Read Detection of RNAi-Induced Protein Knockdown in Mammalian Cells by Western Blotting Protocol]

Category: Reporter Gene Assays

Protocol for GUS reporter gene assay. Includes: Protein isolation; Alternative method for small (<1g) quantities of tissue; GUS assays; Bradford protein concentration determination assays - [Read GUS Reporter Gene Assay Protocol]

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to distinguish a Spore killer from alternative causes of ascospore abortion. - [Read How to Distinguish a Spore Killer From Alternative Causes of Ascospore Abortion]

Category: Immunology: Immunofluorescence Protocols

This protocol describes the use of a specific antibody that recognizes the targeted gene product to detect RNAi-induced gene knockdown in mammalian cells. Western blot technology can be used as an alternative (see Detection of RNAi-Induced Protein Knockdown in Mammalian Cells by Western Blotting). - [Read Immunofluorescence Detection of RNAi-Induced Protein Knockdown in Mammalian Cells Protocol]

Category: RNAi Technology Protocols: RNAi Mammalian Cells Protocols

Protocol describes the use of a specific antibody that recognizes the targeted gene product to detect RNAi-induced gene knockdown in mammalian cells.  Western blot technology can be used as an alternative. - [Read Immunofluorescence Detection of RNAi-Induced Protein Knockdown in Mammalian Cells Protocol]

Category: C. elegans Protocols: C. elegans Genetics Protocols

Protocol describes an easily scalable way of introducing double-stranded RNA (dsRNA) in Caenorhabditis elegans: feeding the nematode with bacteria that express dsRNA. When using an RNase-III-negative Escherichia coli strain (HT115), the efficiency of this method is comparable to the alternative. - [Read Introduction of Double-Stranded RNA in C. elegans by Feeding Protocol]

Category: RNAi Technology Protocols: RNAi C. elegans Protocols

Protocol describes an easily scalable way of introducing double-stranded RNA (dsRNA) in Caenorhabditis elegans: feeding the nematode with bacteria that express dsRNA.  When using an Rnase-III-negative Escherichia coli strain (HT115), the efficiency of this method is comparable to the alternative. - [Read Introduction of Double-Stranded RNA in C. elegans by Feeding Protocol]

Category: C. elegans Protocols: C. elegans Genetics Protocols

Double-stranded RNA (dsRNA) can be introduced into Caenorhabditis elegans by soaking the animals in a solution of dsRNA. Alternative methods are dsRNA injection (see Introduction of Double-stranded RNA in C. elegans by Injection) and feeding the animals with bacteria that produce dsRNA. - [Read Introduction of Double-Stranded RNA in C. elegans by Soaking Protocol]

Category: RNAi Technology Protocols: RNAi C. elegans Protocols

Double-stranded RNA (dsRNA) can be introduced into Caenorhabditis elegans by soaking the animals in a solution of dsRNA.  Alternative methods are dsRNA injection and feeding the animals with bacteria that produce dsRNA. - [Read Introduction of Double-Stranded RNA in C. elegans by Soaking Protocol]

Category: Cell Biology and Cell Culture: Cell Viability Protocols

Assay measures cell viability. It is a two-color fluorescence assay that simultaneously determines Live cell number and Dead cell number. This protocol is designed for use with the GEMINI XS Microplate Spectrofluorometer, a multi-well plate scanner with dual excitation/emission capabilities, but the assay is also adaptable for flow cytometry and fluorescence microscopy. Includes: Cell Culture; Preparation for the Assay; Live/Dead Assay; Reading the Plate; Data Analysis; Alternative protocol. - [Read Live/Dead Assay for Cell Viability Protoco]

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]

Category: Microsphere Analysis Protocols

The role of microspheres in these screens is similar to their traditional role in immunoassays, namely as a solid phase to either enhance detection, separation, or both. The predominance of radioactive assays in high-throughput screening, along with the desire to find alternative means of detection, have led to research on substituting alternative fluorescent technologies. - [Read Microspheres for High-Throughput Screening Assays]

Category: Microscopy Protocols: Optical Microscopy Protocols

Near-field scanning optical microscopy can achieve spatial resolution performance beyond the classical diffraction limit by employing a sub-wavelength light source or detector positioned in close proximity to a specimen. Such a sub-wavelength source usually consists of an aperture at the end of a tapered probe, which functions basically as a wave guide. Includes info.: Fiber Probe Fabrication; Pulling Method; Meniscus Etching; Selective Etching; Apertureless and Alternative Probe Designs etc. - [Read Near-Field Scanning Optical Microscopy: NSOM Probes]

Category: Yeast Cell Culture Protocols: Yeast Sporulation Protocols

Zygotes can be identified by their unique morphology. They can be easily separated away from nonmated cells using a micromanipulator. This method provides an alternative to the selection of diploid cells on a medium that prevents the growth of haploid parent cells. - [Read Picking Zygotes Protocol]

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]

Category: Chromatography Protocols: Dye Ligand Affinity Chromatography Protocols

Triazine dyes, such as Cibacron Blue 3GA, can be linked to a hexyl spacer arm and then immobilized on a polyhydroxyl support matrix that has been activated with either 1,1-carbonyldiimidazole or epichlorohydrin.  An alternative procedure for immobilizing dyes using the direct coupling method is provided in Immobilization of Dyes on Polyhydroxyl Matrices Using the Direct Coupling Method. - [Read Preparation of Affinity-Ligand Resins by Immobilization of Dyes on Polyhydroxyl Matrices]

Category: Immunology: B Cell Protocols

Describes procedures for measuring the capacity of purified B cells to undergo proliferation. The method centers on the use of polyclonal stimulating agents (mitogens) because these agents stimulate the majority of B cells and because the alternative (measurement of antigen-induced proliferation) requires the laborious procedures of isolating antigen-specific B cells (which are otherwise present in too low a concentration in whole B cell populations). - [Read Proliferative Assays for B Cell Function Protocol]