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(LCM): Preparation and Sectioning of Frozen Tissue Blocks and Purification of RNA from Isolated Cel - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4107

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]

96-Well Confirmation Yeast PCR Protocol - http://sequence-www.stanford.edu/group/yeast_deletion_project/96_well_confirmation.html

Category: Yeast Protocols: Yeast Nucleic Acid Protocols: Yeast PCR Protocols

Protocol for 96-well confirmation Yeast PCR. Includes: Clonal purification; Generate a master plate (96-well format); Making a frozen backup stock; Confirmation PCR for one Row; ORF Specific Confirmation PCR --> "A-B" primers (upstream junction); Transfer template DNA to multiwell PCR plate; Prepare and dispense master mix for A-B PCR. - [Read 96-Well Confirmation Yeast PCR Protocol]

ANTIBODY PURIFICATION by affinity chromatography - http://mullinslab.ucsf.edu/protocols/html/ANTIBODY_PURIFICATION_by_affini.htm

Category: Chromatography Protocols: Immunoaffinity Chromatography Protocols

ANTIBODY PURIFICATION by affinity chromatography. By Beth, Mullins Lab UCSF. To affinity purify antibodies, generate lots of E. coli lysate that contains your antigen. If the protein can stand freeze thawing, then go ahead and purify the protein from e. coli lysate and keep it frozen until you need to couple it to a CH-sepharose column. - [Read ANTIBODY PURIFICATION by affinity chromatography]

Basic Techniques for Mammalian Cell Tissue Culture Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E662B5FE053DEE8CB4AC3EB6F729A11&objectid=667396C6F0D96C40EB68A3D8DB8B201A

Category: Cell Culture Protocols: Sterile Technique Protocols

Cultured mammalian cells are used extensively in cell biology studies; it requires a number of special skills in order to be able to preserve the structure, function, behavior and biology of the cells. This unit describes the basic skills required to maintain and preserve cell cultures: aseptic technique, medium characteristics, passaging, freezing and storage, recovering frozen stocks, and counting viable cells. - [Read Basic Techniques for Mammalian Cell Tissue Culture Protocol]

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

cDNA from Frozen Tissue Sections and LCM - http://dir.nichd.nih.gov/lcm/prep.htm

Category: RNA Protocols: RT-PCR and cDNA synthesis

Method of synthesizing cDNA from Frozen Tissue Sections Protocol. Includes LCM - Laser Capture Microdissection Protocols. NIH - [Read cDNA from Frozen Tissue Sections and LCM]

cDNA Synthesis and Real-Time PCR Using RNA from Laser-Captured Cells Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4108

Category: PCR Protocols: Real-Time PCR Protocols

Protocol describes the real-time PCR using RNA purified from laser-captured tissues Laser Capture Microdissection (LCM): Preparation and Sectioning of Frozen Tissue Blocks and Purification of RNA from Isolated Cells. - [Read cDNA Synthesis and Real-Time PCR Using RNA from Laser-Captured Cells Protocol]

CHO Centrosome Prep Protocol - http://mitchison.med.harvard.edu/protocols/mt1.html

Category: Cell Organelle Protocols: Nucleus Protocols

The key step is the lysis which solubilizes centrosomes away from nuclei by very low ionic strength lysis after treatment of cells with nocodazole and cytochalasin B. The released centrosomes are then centrifuged onto a Ficoll cushion (to avoid pelleting) and the interface between the lysate and the Ficoll is collected and the centrosomes are concentrated on a sucrose gradient. Fractions are assayed by spindown and double IF with 5051 serum and anti-tubulin and the pooled fractions are frozen... - [Read CHO Centrosome Prep Protocol]

Clonality - X Chromosome Inactivation Assay Protocol - http://cgap-mf.nih.gov/Protocols/DNARNAProteomicAnalysis/DNA/DNAProtocols/Clonality.html

Category: Genetics and Genomics: Cancer Genetics Protocols

Investigators can utilize X chromosome inactivation (methylation) to determine the clonality status of a tumor or premalignant lesion in females. The technique is based on a methylation-sensitive restriction enzyme and analysis of a polymorphic locus on the X chromosome. Clonal cell populations will show "loss" of the non-methylated allele after restriction digest. The assay can be performed on DNA recovered from microdissected samples. Both frozen tissue and fixed-embedded tissue can be used. - [Read Clonality - X Chromosome Inactivation Assay Protocol]

Competent cell preparation - http://www.chem.uga.edu/scottgrp/GrpProtocols/Competent_cell_preparation.htm

Category: Microbiology: Bacterial Competent Cells Preparation E.Coli

Bacterial E.Coli competent cell preparation. Calcium Chloride Protocol, Preparation of calcium chloride competent cells for frozen storage, Electroporation Protocol, Electroporation Protocol for transformations using double-stranded plasmids, Reference: - [Read Competent cell preparation]

Cryopreservation of Mammalian Culture Cells: Preparation and Recovery of Samples Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4350

Category: Cell Culture Protocols: Cell Storing and Cell Freezing Protocols

Protocol provides methods for cryofreezing and subsequent thawing of mammalian cells. Pre-confluent cells are trypsinized, pelleted, resuspended in freezing medium, and gradually frozen. When needed, frozen cells are thawed quickly under running tap water and transferred to growth medium. - [Read Cryopreservation of Mammalian Culture Cells: Preparation and Recovery of Samples Protocol]

Culture of Human Prostatic Carcinoma Cell Lines - http://www.celldeath.de/apometh/rpmi.html

Category: Cell Biology and Cell Culture: Cell Line Culture Conditions

Culture of Human Prostatic Carcinoma Cell Lines, Growing and splitting the cells, Preparation of frozen stocks in liquid nitrogen, How to bring frozen cells back into culture, Concentrations of antibiotics for the selection of stable transfectants. LNCa - [Read Culture of Human Prostatic Carcinoma Cell Lines]

Defrosting Eukaryotic Cells Frozen In Liquid N2 Protocol - http://www2.umdnj.edu/~geller/lab/thawing.html

Category: Cell Culture Protocols: Cell Storing and Cell Freezing Protocols

Protocol for the defrosting of eukaryotic cells frozen in liquid N2. - [Read Defrosting Eukaryotic Cells Frozen In Liquid N2 Protocol]

Frozen Competent E. coli Preparation and Transformation Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/F3.html

Category: E Coli Protocols: E coli Transformation Protocols

Protocol for frozen competent E. coli preparation and transformation. - [Read Frozen Competent E. coli Preparation and Transformation Protocol]

Frozen Competent E. coli Preparation and Transformation Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/F3.html

Category: Competent Cells Protocols

This is a very easy protocol which yields transformation efficiencies appropriate for routine cloning and transformation. - [Read Frozen Competent E. coli Preparation and Transformation Protocol]

Frozen Competent E.Coli Cells - http://www.ciwemb.edu/labs/koshland/Protocols/BACTERIA/ecolicells.html

Category: Microbiology: Bacterial Competent Cells Preparation E.Coli

Frozen Competent E.Coli Cells. Methods and protocol for freezing E.Coli Bacteria Competent Cells in aliquots for later use re-use. (Inoue et al., 1990 Gene 96:23). Koshland. - [Read Frozen Competent E.Coli Cells]

Frozen Sectioning and Tissue Sectioning for Electron Microscopy - http://www.ucsf.edu/pathol/research/morphprotocol.htm

Category: Histology Protocols: Tissue Sample Sectioning

Frozen Sectioning and Tissue Sectioning for Electron Microscopy. Includes Tissue Processing for Transmission Electron Microscopy and Tissue Preservation for Light Microscopy. University of California UCSF Dept. Pathology Research. - [Read Frozen Sectioning and Tissue Sectioning for Electron Microscopy]

FROZEN SECTIONING PROTOCOL Plant Tissue PDF - http://schnablelab.plantgenomics.iastate.edu/docs/resources/protocols/Cryosectioning_for_LCM.pdf

Category: Histology Protocols: Tissue Sample Sectioning

PREPARATION OF PLANT TISSUE FOR LASER MICRODISSECTION. Schnable Labs, Plant Genomics. - [Read FROZEN SECTIONING PROTOCOL Plant Tissue PDF]

Frozen Yeast Transformation Protocol - http://www.umanitoba.ca/faculties/medicine/biochem/gietz/Frozen.html

Category: Yeast Protocols

This Protocol allows You to prepare Frozen Yeast cells that are competent for transformation after thawing. - [Read Frozen Yeast Transformation Protocol]

Genetic Mapping of Arabidopsis Centromeres Using PCR-Based Markers Protocol - http://www.arabidopsis.org/cshl-course/3-genetic_mapping.html

Category: Plant Biology Protocols: Arabidopsis Genetics Protocols

Protocol for genetic mapping of Arabidopsis centromeres using PCR-based markers. Includes: DNA markers facilitate genome analysis; Centromere mapping using tetrad analysis; Preparation of DNA from frozen plant tissue. - [Read Genetic Mapping of Arabidopsis Centromeres Using PCR-Based Markers Protocol]

Genome-Wide Analysis of Protein-Protein Interactions Using a Two-Hybrid Array: S - http://www.cshprotocols.org/cgi/content/abstract/2006/23/pdb.prot4564

Category: Yeast Protocols: Yeast Protein Protocols: Yeast Protein Protein Interactions Protocols

This protocol describes three different storage methods for the array. Frozen stocks are the most permanent method and are an absolute requirement for maintaining a record of the array. - [Read Genome-Wide Analysis of Protein-Protein Interactions Using a Two-Hybrid Array: S]

HISTOLOGY-RELATED PROTOCOLS Fixation and Frozen Block PDF - http://www.uchsc.edu/misc/diabetes/derc/protocols/histologyprotocols.pdf

Category: Histology Protocols: Fixation of Cells Tissues Protocol

Protocols for cell fixation and tissue fixation. HISTOLOGY-RELATED PROTOCOLS Fixation and Frozen Block techniques. Embedding Tissue Into Frozen Blocks. Fixing Cells with Fresh Paraformaldehyde. Preparation of Frozen Sections. Immunohistochemistry. Paraffin sections and Frozen sections. DERC Univ. Colorado PDF. - [Read HISTOLOGY-RELATED PROTOCOLS Fixation and Frozen Block PDF]

Histotechniques Fixation - http://library.med.utah.edu/WebPath/HISTHTML/HISTOTCH/HISTOTCH.html

Category: Immunohistochemistry Protocols: Fixation Protocols

Information on histotechniques. Includes: Fixation - types of fixatives; factors affecting fixation; General usage of fixatives; Tissue Processing; Sectioning; Frozen Sections; Staining; H and E staining; Coverslipping; Decalcification; Artefacts in Histologic Sections; Problems in Tissue Processing. - [Read Histotechniques Fixation]

Immuno-Laser Capture Microdissection Protocol - http://cgap-mf.nih.gov/Protocols/Slides/SlideProtocols/ImmunoLCM.html

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

After fixation, frozen sections are immunostained under RNase-free conditions using a rapid three-step streptavidin-biotin technique followed by dehydration. The immunostained sections are ready for LCM. Includes: Development of Immuno-LCM. - [Read Immuno-Laser Capture Microdissection Protocol]

Immunofluorescence Staining Protocol - http://www.ihcworld.com/_protocols/general_IHC/immunofl.htm

Category: Immunology: Immunofluorescence Protocols

Protocol for immunofluorescence staining. Includes: Cell Lines; Frozen Sections; Paraffin Sections; Pretreatments of Tissue Sections. - [Read Immunofluorescence Staining Protocol]

Articles (1-2 of 2)

Immobilized Antigen Phage Antibody Selection Protocol

A protocol for the selection of Phage Antibodies using Immobilized Antigen. This method describes the selection of antibodies from bacteriophage antibody libraries that recognize a specific antigen. The phage display library of antibody-displaying phage particles is exposed to antigen attached to a solid substrate (Nunc Immuno™ tubes). The phage particles with affinity for antigen bind to the immobilized antigen and are selected from the library of phage expressing antibodies.