part Protocols

Category: PCR Protocols: cDNA Synthesis Protocols

To generate "3'-end" partial cDNA clones, mRNA is reverse-transcribed using a "hybrid" primer (Qtotal, QT) that consists of two mixed bases (GATC/GAC followed by [T]17) and a unique 35-base oligonucleotide sequence (QI-QO).  Amplification is then performed using a primer containing part of this sequence (Qouter, Qo) (which now binds to each cDNA at its 3'-end) and a primer derived from the gene of interest, GSP1 (gene-specific primer 1). - [Read 3'-End cDNA Amplification Using Classic RACE Protocol]

Category: Cell Biology and Cell Culture: ES Stem Cell Biology and Culture: MEF Protocols Mouse Embryonic Fibroblasts

WiCell Research Institute. Recipes, Frequently Asked Questions, Mouse Embryo Extraction and Tissue, Harvesting Mouse Embryonic Fibroblasts (MEF)from mice. - [Read Derivation of Mouse Embryonic Fibroblasts (MEF) Part I PDF]

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]

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

Development of techniques for primary culture of C. elegans embryonic neurons. - [Read Development of Techniques for Primary Culture of C. elegans Embryonic Neurons 3rd Part]

Category: Microscopy Protocols: Electron Microscopy Protocols

Article describe the preparation of cells for correlative electron microscopy after live light microscopic observation of fluorescently labeled cytoskeletal proteins microinjected into the same cells. Since identification of cytoskeletal elements in electron microscopic preparations is an essential part of any correlative study, procedures for immunogold labeling of cytoskeletal components and for myosin S1 decoration of actin filaments are also described. - [Read Electron Microscopy of the Cytoskeleton of Cultured Cells]

Category: Genetics and Genomics: Genotyping Protocols: PCR Genotyping

This protocol describes a rapid PCR-based method for identifying targeted ES cell colonies prior to picking. It is based on DNA analysis of a small part of colonies pooled directly from selection plates. Only positive colonies are expanded. - [Read Genotyping Embryonic Stem (ES) Cell Colonies Prior to Picking Protocol]

Category: Cell Biology and Cell Culture: ES Stem Cell Biology and Culture: MEF Protocols Mouse Embryonic Fibroblasts

WiCell Reserarch Institute. Count MEF Cells, Trypsinize, Split, Freeze, Thaw, Irradiate, Reconstitute, Plate, Feeder. - [Read Mouse Embryonic Fibroblast Culture and Propagation Part II PDF]

Category: Pharmacology and Toxicology Protocols: Pharmacokinetics Protocols

The purpose of the present protocol is to obtain this information through collaboration with interested physicians and patients taking part in the European AED Pregnancy Registry (EURAP). - [Read Pharmacokinetic Evaluation of Antiepileptic Drugs During Pregnancy]

Category: Protein Protocols: Protein Precipitation Procols

Laboratory sample cleanup is a necessary part for analytical preparation analysis. The removal of Contaminants such as proteins, cell debris and other materials is an important step. Typically this has been done by using Acetonitrile and then Centrifugation to pellet the debris leaving the clean supernant. After this process supernatant can be used for further analysis by HPLC, GC, MS and other analysis tandem methods. HTS Labs. - [Read Protein Precipitation Microplate]

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

The first part of the isolation procedure is a flotation through a continuous iodixanol gradient; this gradient is essentially a resolving gradient in which the caveolin-rich vesicles are concentrated in the top third of the gradient, while the predominantly caveolin-poor vesicles band in denser regions. A second discontinuous gradient is essentially a concentration gradient to band the caveolin-rich vesicles sharply at an interface. - [Read Purification of Caveolae Membranes from a Plasma Membrane Fraction of Cultured Cells and Tissues]

Category: PCR Protocols: Reverse Transcriptase RT-PCR Protocols

In the first part of this protocol, the linear range of amplification is determined by carrying out 10 identical PCRs in the presence of [{alpha}-32P]dCTP and stopping one reaction after every two cycles.  Amplification products are quantified on a denaturing polyacrylamide gel and the results plotted on a graph (counts per minute vs. cycle number).  Total RNA is used as an internal control. - [Read Relative RT-PCR: Determining the Linear Range of Amplification and Optimizing the Primers:Competimer]