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Calcium-phosphate-mediated Transfection of Eukaryotic Cells with Plasmid DNAs - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3871

Category: Transfection Protocols: Calcium Phosphate Transfection Protocols

Calcium phosphate forms an insoluble precipitate with DNA, which attaches to the cell surface and is taken into the cells by endocytosis. The protocol is easily adapted for use with other types of cells, both adherent and nonadherent. This protocol is a modified version of a method published by Jordan et al. (1996) who rigorously optimized calcium-phosphate-based transfection methods for Chinese hamster ovary cells and the 293 line of human embryonic kidney cells. - [Read Calcium-phosphate-mediated Transfection of Eukaryotic Cells with Plasmid DNAs]

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]

Detection of mRNAs on Cryosections of the Cardiovascular System Using DIG-Labeled RNA Probes - http://www.roche-applied-science.com/PROD_INF/MANUALS/InSitu/pdf/ISH_189-196.pdf

Category: Cytogenetics Protocols: In Situ Hybridization Tissue Sections Protocols

Protocol for detection of mRNAs on cryosections of the cardiovascular system using DIG-labeled RNA probes. Protocol was optimized from a protocol using 35S-labeled RNA probes. It allows to detect the expression of low abundant mRNAs in the cardiovascular system, e.g. of the proinflammatory cytokine GM-CSF in normal human coronary arteries, and of IL6 and gp130 in human failing hearts. The protocol can be combined with immunohistochemistry. - [Read Detection of mRNAs on Cryosections of the Cardiovascular System Using DIG-Labeled RNA Probes]

Development of Techniques for Primary Culture of C. elegans Embryonic Neurons 2nd Part - http://cobweb.dartmouth.edu/cgi-bin/cgiwrap/~ambros/protocol.cgi?id=15

Category: C. elegans Protocols: C. elegans Cell Culture Protocols

Development of techniques for primary culture of C. elegans embryonic neurons. Includes: Optimized cell culture protocol. - [Read Development of Techniques for Primary Culture of C. elegans Embryonic Neurons 2nd Part]

Electro Mobility Shift Assay Protocol - http://ruummc.med.uu.nl/subsite/RepHTML/Protocols/EMSARdJ.html

Category: DNA Protein Interactions Protocols: Electrophoretic Mobility Shift Assays EMSA Protocols

Protocol electro mobility shift assay (EMSA) optimized for pTP. - [Read Electro Mobility Shift Assay Protocol]

Extraction and Solubilization of Total Protein from Microorganisms Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4224

Category: Bacterial Cell Culture Protocols

The growth conditions of microbial cell cultures and the time of sample collection should be optimized and standardized when growing cells for protein extraction. Because cells may excrete proteases and other extracellular enzymes, and compounds in the medium may interfere with extraction, wash cultures with an isotonic buffer, such as PBS or sucrose before solubilization. - [Read Extraction and Solubilization of Total Protein from Microorganisms Protocol]

Histochemical Staining Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/H2.html

Category: Immunohistochemistry Protocols: Antibody Staining Protocols

Protocol for histochemical staining. This protocol has been optimized for b-galactosidase and human placental alkaline phosphatase staining in retinal tissue and cultured cells. - [Read Histochemical Staining Protocol]

Immunoperoxidase Methods for Localization of Antigens in Cultured Cells and Tissues - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E662E8B08D36AFBDC097EF76E740CB1&objectid=6673CDF3E6A07EBE28C6002718B3CC7A

Category: Immunohistochemistry Protocols

Describes two methods for using the immunoperoxidase reaction to localize antigens at the electron microscope level; one for adherent cultured cells and one for tissue sections. The reaction conditions are first optimized at the light microscope level and then adapted for EM level observation. These methods allow for reliable detection of antigens at the cell surface, within the cell, and especially in membrane bounded organelles. - [Read Immunoperoxidase Methods for Localization of Antigens in Cultured Cells and Tissues]

Optimized Protocol for Mounting Tissue Sections PDF - http://arctur.com/research_portal/images/Application%20Notes/pen_membrane_protocol_9.pdf

Category: Histology Protocols: Tissue Sample Sectioning

Generally Required Materials and Equipment for Tissue Sectioning. Sectioning and Slide Preparation Procedure onto Metal-Framed PEN Membrane Slides. Arcturus. - [Read Optimized Protocol for Mounting Tissue Sections PDF]

Optimized Protocols for Fluorescent in situ Hybridization in Drosophila Tissues - http://www.utoronto.ca/krause/documents/FISHCHAPTER.pdf

Category: Drosophila Protocols: Drosophila Genetics Protocols

Optimized protocols for fluorescent in situ hybridization in Drosophila tissues. Includes: RNA Probe Preparation; Initial Embryo Fixation; Post-Fixation, Hybridization and post-Hybridization Washes; Development of FISH Signal; Mounting and Viewing of Samples; Double FISH; FISH on Dissected Tissues; RNA-Protein Double-labeling. - [Read Optimized Protocols for Fluorescent in situ Hybridization in Drosophila Tissues]

Optimized Welsh DDRT-PCR Protocol - http://wheat.pw.usda.gov/~lazo/methods/lazo/ddrtpcr2.html

Category: DNA Protocols: PCR Polymerase Chain Reaction Protocols

Optimized Welsh DDRT-PCR Protocol. Provided by Mirta Grifman and found in Neuroscience Protocols (Nov 1995). - [Read Optimized Welsh DDRT-PCR Protocol]

Optimized Welsh DDRT-PCR Protocol - http://wheat.pw.usda.gov/~lazo/methods/lazo/ddrtpcr2.html

Category: PCR Protocols

Optimized Welsh DDRT-PCR Protocol. Provided by Mirta Grifman and found in Neuroscience Protocols (Nov 1995). - [Read Optimized Welsh DDRT-PCR Protocol]

Optimized Welsh DDRT-PCR Protocol - http://wheat.pw.usda.gov/~lazo/methods/lazo/ddrtpcr2.html

Category: PCR Protocols: Reverse Transcriptase RT-PCR Protocols

Protocol for optimized welsh DDRT-PCR. - [Read Optimized Welsh DDRT-PCR Protocol]

Peroxisomes in Saccharomyces cerevisiae - http://mcb.asm.org/cgi/reprint/11/1/510.pdf

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Peroxisome Isolation Protocols

To isolate peroxisomes from Saccharomyces cerevisiae of a quality sufficient for in vitro import studies, we optimized the conditions for cell growth and for cell fractionation. Stability of the isolated peroxisomes was monitored by catalase latency and sedimentability of marker enzymes. - [Read Peroxisomes in Saccharomyces cerevisiae]

Preparation of Nuclear Extracts for Gel Shifts and Westerns Blots - http://faculty.virginia.edu/mirmira/resources_files/Protocols/Nuclear%20Extracts.htm

Category: Protein Protocols: Nuclear Extraction Protocols

Nuclear Extract protocol is optimized for about 10^7 cells, can use for gel shift assays and western blotting. Mirmira Lab, Univ. Virginia - [Read Preparation of Nuclear Extracts for Gel Shifts and Westerns Blots]

Propagation of Pluripotent Mouse Embryonic Stem (ES) Cells Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4484

Category: Stem Cell Protocols: Feeder Cell Preparation Protocols

This protocol describes the culture of embryonic stem (ES) cells using mitotically inactivating primary mouse embryonic fibroblast (MEF) cells as a feeder layer (preparation described in Preparation of Mouse Embryonic Fibroblast (MEF) Feeder Plates). The ES culture medium is supplemented with recombinant leukemia inhibitory factor (LIF) to help maintain the cells as pluripotent stem cells. This protocol has been optimized for the ES-D3 cell line. - [Read Propagation of Pluripotent Mouse Embryonic Stem (ES) Cells Protocol]

Propagation of Pluripotent Mouse Embryonic Stem (ES) Cells Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/18/pdb.prot4484

Category: Stem Cell Protocols: Mouse Embryonic Fibroblasts Protocols

Protein Purification Using Immobilized Metal-Ion Affinity Chromatography under Optimized Imidazole - http://www.cshprotocols.org/cgi/content/extract/2006/9/pdb.prot4262

Category: Chromatography Protocols: Immobilized Metal Ion Affinity Chromatography Protocols

Perform this protocol after determining the imidazole concentration that yields the best purification of the recombinant protein of interest. - [Read Protein Purification Using Immobilized Metal-Ion Affinity Chromatography under Optimized Imidazole]

Protocol for Stable Lines - http://genetics.med.harvard.edu/~cepko/protocol/mike/S4.html

Category: Primary Cell Isolation and Culture Protocols: Mammalian Cell Culture Protocols

This protocol is optimized for erythroid cells, other cell types may require modifications. - [Read Protocol for Stable Lines]

Purification of Histidine-Tagged Proteins Using IMAC Without Parameter Optimization Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4217

Category: Chromatography Protocols: Immobilized Metal Ion Affinity Chromatography Protocols

Histidine-tagged proteins can be purified on prepacked 1-ml immobilized metal-ion affinity chromatography (IMAC) columns without optimization of the separation conditions. The method allows fast capture of the target protein, although with a lower purity than can be obtained under optimized conditions. - [Read Purification of Histidine-Tagged Proteins Using IMAC Without Parameter Optimization Protocol]

Rat Neuron Nucleofector Kit - http://www.amaxa.com/fileadmin/groups/marketing/Downloads/Protocols/Primary_cells/amaxa_OP_Neuron_hippo_cortical_rat_DPG-1003.pdf

Category: Neuroscience Protocols: Neuronal Cell Culture Protocols: Rat Neuronal Cell Culture Protocols

Optimized Protocol for primary rat hippocampal or cortical neurons. - [Read Rat Neuron Nucleofector Kit]

The Procedure for Optimized ELISA Protocol - http://www.pall.com/34445_4769.asp

Category: ELISA Protocols

The Procedure for Optimized ELISA Protocol. PALL Corp. - [Read The Procedure for Optimized ELISA Protocol]

Transfection of Hippocampal Neurons with Plasmid DNA Using Calcium Phosphate Coprecipitation - http://www.cshprotocols.org/cgi/content/abstract/2006/1/pdb.prot4445

Category: Transfection Protocols: Calcium Phosphate Transfection Protocols

This protocol describes transfection of plasmid DNA into primary hippocampal neurons using DNA/calcium-phosphate (CaPO4) coprecipitation. The precise pH of the transfection medium and the incubation time of cells with the coprecipitate are critical for reproducible and efficient transfection. Once these parameters are optimized for a given plasmid, the method is easily adapted for transfection of other established cell lines. - [Read Transfection of Hippocampal Neurons with Plasmid DNA Using Calcium Phosphate Coprecipitation]

Transient Transfection of Cos-1 Cells - http://www.upstate.com/misc/protocols.q.prot.e.cDNAprotocol/Transient+Transfection+of+Cos+1+Cells

Category: Cell Biology and Cell Culture: Cell Transfection Protocol

Protocol optimized for Cos-1 cells. Successful transfection of each cell type requires optimization of the basic protocol. LIPOFECTAMINE method of transient transfection. Upstate. - [Read Transient Transfection of Cos-1 Cells]

Use of PCR for Quality Control of a Genomic DNA Library Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4138

Category: DNA Protocols: Genomic DNA Library Protocols

Protocol describes the use of PCR to analyze the quality of a human genomic DNA library. It has been optimized for a human genomic DNA library containing fragments between approximately 50 and 500 bp. - [Read Use of PCR for Quality Control of a Genomic DNA Library Protocol]