tissue Protocols

Category: Immunohistochemistry Protocols: Antibody Staining Protocols

Staining of tissues can be carried out on cell smears prepared from biopsy samples, such as needle aspirates, tissue scrapings, or freshly dissected tissues. - [Read Preparing Cell Smears from Tissue Samples for Immunostaining Protocol]

Category: Immunohistochemistry Protocols: Antibody Staining Protocols

Frozen tissue sections show good preservation of tissue structure and antigens. The principle disadvantages of using them in immunostaining are that the specimens must be stored frozen, and a special microtome, known as a cryostat, is required. Also, many clinical specimens are not available in this form, and most classic histological descriptions of tissue structure and pathology are based on the use of paraffin-embedded sections of formalin-fixed material. - [Read Preparing Frozen Tissue Sections for Immunostaining Protocol]

Category: Immunohistochemistry Protocols: Antibody Staining Protocols

Most histological studies are carried out on paraformaldehyde-fixed, paraffin-embedded tissue samples. Therefore, there is an extensive atlas of most tissues and organs prepared from these sources, and comparing the location of antigens to these data is immediately informative. The fixation and embedding procedures are harsh, however, and many antigens are not well preserved. - [Read Preparing Paraffin Tissue Sections for Immunostaining Protocol]

Category: Radioactivity: Liquid Scintillation Counting

Excellent guide for Liquid Scintillation Counting. Includes protocols and methods for counting gel slices, SPECIAL SAMPLE PREPARATION PROTOCOLS: TLC Plates, protocol for Counting Samples on Cellulose-ester, Filters (MilliporeTM filters), Counting Tissue radioacitivity, Counting 14CO2, Samples in Polyacrylamide Gels. National Diagnostics. National Diagnostics Laboratory Staff. - [Read Principles and Applications of Liquid Scintillation Counting PDF]

Category: Histology Protocols: Fixation of Cells Tissues Protocol

Cell Fixation. 1. Gently wash cells 2x in sterile 1XPBS, ... [Take to Research Histology for embedding] Instructions: Embed pellets in vertical ... Jena Giltnane Yale University Dept. of Pathology. Processing Adherent Cultured Cells for Tissue Microarray Controls. - [Read Processing Adherent Cultured Cells for Tissue Microarray Controls ...]

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

Information on processing of microdissected tissue for molecular analysis. Includes: More than 10,000 Cells; Less than 10,000 Cells; Formalin-fixed; Paraffin-embedded Tissue; Other Fixatives; Timing. - [Read Processing of Microdissected Tissue for Molecular Analysis]

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

The methodology and buffers utilized for processing microdissected samples for protein-based studies is variable depending on the downstream molecular analysis method. - [Read Processing of Microdissected Tissue for Molecular Analysis Proteomics-based Studies]

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

RNA-based studies of processing of microdissected tissue for molecular analysis. Includes: Total RNA Isolation; Optional DNAse Treatment. - [Read Processing of Microdissected Tissue for Molecular Analysis RNA-based Studies]

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: DNA Staining Protocols for Flow Cytometry

This protocol describes a method for quantitative measurement of DNA in solid tissue samples using either propidium iodide (PI) or DAPI staining followed by flow cytometry. PI can be excited at 488 nm by the argon-ion laser, the most commonly used laser in flow cytometry. Alternatively, DAPI is best excited by a high-power UV laser, which is less commonly available. - [Read Propidium Iodide (PI) or DAPI Staining of Unfixed Solid Tissues for Flow Cytometry Protocol]

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: DNA Staining Protocols for Flow Cytometry

This protocol describes a method for quantitative measurement of DNA in tissue culture cells using either propidium iodide (PI) or DAPI staining followed by flow cytometry. PI can be excited at 488 nm by the argon-ion laser, the most commonly used laser in flow cytometry. Alternatively, DAPI is best excited by a high-power UV laser, which is less commonly available. - [Read Propidium Iodide (PI) or DAPI Staining of Unfixed Tissue Culture Cells for Flow Cytometry Protocol]

Category: Protein Protocols: Protein Extraction From Tissue

Combination of nucleic acid and protein isolation with tissue array construction: Using defined histologic regions in single frozen tissue blocks for multiple research purposes - [Read Protein isolation with tissue array construction]

Category: Transfection Protocols: Electroporation Sonoporation Protocols

Protocol describes a method for introducing gene constructs into mouse embryos by electroporation. Gene constructs can be quickly tested for tissue-specific transcriptional activity or can be used to overexpress gene products. - [Read Protocol Electroporation]

Category: Signalling Signal Transduction Protocols

Protocol describes a competitive ligand binding assay for cortical neurotrophin receptors. Following binding in the presence of competitor, the bound radiolabeled ligand is cross-linked to the receptor. The cells are lysed and the ligand-receptor complexes are immunoprecipitated using a pan-trk (tyrosine kinase receptor) antibody. Protocol includes:Preparation of Cortical Tissue for Competitive Crosslinking, Competitive Binding, Crosslinking Ligand to Receptor, Lysis and Immunoprecipitation etc - [Read Protocol for Competitive Ligand Binding to Cortical Receptor using Crosslinking]

Category: Immunology: Immunoprecipitation Protocols

Immunoprecopitation method, the protein from the cell or tissue homogenate is precipitated in an appropriate lysis buffer by means of an immune complex which includes the antigen (protein), primary antibody and Protein A-, G-, or L-agarose conjugate or a secondary antibody-agarose conjugate - [Read Protocol Immunoprecipitation]

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

Protocols for LCM preparation and analysis. Includes protocols: Preparation, LCM and RNA/DNA extraction of Frozen Tissue Sections; Preparation and LCM of Paraffin Embedded Tissue Sections; Standard Protocols for Microdissected Tissue Analysis. - [Read Protocols for LCM Preparation and Analysis]

Category: Cytoskeleton Protocols: Intermediate Filaments Protocols

Intermediate filaments (IF) are major cytoskeletal systems of vertebrate and many nonvertebrate cells whose expression is cell-type specific and developmentally regulated. This protocol describes a method for purifying one type of IF, vimentin, from bovine lens tissue. Purification of human vimentin expressed in Escherichia coli is also described. These methods are useful in the preparation of other IF protein subunits for microinjection studies as well. - [Read Purification of Bovine Lens and Bacterially Expressed Human Vimentin Protocol]

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols: RNA Isolation fromTissues or Cultured Cells Protocols

Single-step technique, cells are homogenized in guanidnium thiocyanate and the RNA is purified from the lysate by extraction with phenol:chloroform at reduced pH.  Many samples can be processed simultaneously and speedily.  The yield of total RNA depends on the tissue or cell source and is generally in the range of 4-7 µg/ml starting tissue or 5-10 µg/106 cells.  IMPORTANT: Prepare all reagents used in this protocol with Diethyl pyrocarbonate (DEPC)-treated H2O. - [Read Purification of RNA from Cells and Tissues by Acid Phenol-Guanidinium Thiocyanate-Chloroform Extract]

Category: Plant Biology Protocols: Plant DNA RNA Isolation Protocols

This protocol is not phenol-based, but does require the addition of chloroform. The Concert Plant Reagent is intended for the isolation of RNA from a wide variety of plant tissues including blue spruce needles, potato tuber etc. - [Read Purification of RNA from Plant Tissue Using the Concert Plant Reagent]

Category: Cell Culture Protocols: Cell Culture Contamination Prevention and Removal Protocols

Quality is important in all aspects of tissue culture since the quality of materials used i.e. media and other reagents) will affect the quality of the cultures and products derived from them. The main areas of quality control that are of concern for tissue culture are: The quality of the reagents and materials; The provenance and integrity of the cell lines; The avoidance of microbial contamination. - [Read Quality Control Considerations for Cell Culture]

Category: Plant Biology Protocols: Plant Proteins, Peptides and Antigens

The activity of ß-glucuronidase (GUS) can be accurately determined in intact plant tissue using 4-methylumbelliferyl ß-D-glucuronide (4-MUG) as a substrate. Upon hydrolysis by GUS, the fluorochrome 4-methyl umbelliferone (4-MU) is produced. This method is based on the permeability of both 4-MUG and 4-MU through plant tissue. It consists of incubation of the tissue with the reagent and quantification of the fluorescence emitted by 4-MU in the solution. GUS activity in each sample can be... - [Read Quantitative GUS Activity Assay in Intact Plant Tissue Protocol]

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Isolation from Mammalian Cells Protocols

Mammalian DNA prepared from blood or tissues as described in this protocol is 20-50 kb in size and suitable for use as a template in PCRs.  The yields of DNA vary between 0.5 and 3.0 µg/mg tissue or 5 and 15 µg per 300 µl of whole blood. - [Read Rapid Isolation of Mammalian DNA Protocol]

Category: Stem Cell Protocols

This protocol describes a method for rapid preparation of DNA from cells in 96-well tissue culture dishes. - [Read Rapid Preparation of DNA from Cells in 96-Well Tissue Culture Dishes Protocol]

Category: Mouse Protocols: Mouse Cell Culture Protocols

Protocol for rat Chromaffin cells primary cultures: Standardization and quality assessment for single-cell assays. Includes: Preliminars; Isolation of the rat adrenal medulla; Enzymatic digestion of medullary tissue; Collecting and culturing the cells. - [Read Rat Chromaffin Cells Primary Cultures Protocol]

Category: PCR Protocols: Real-Time PCR Protocols

Protocol uses the Superscript II First-Strand Synthesis system for the generation of cDNA from total RNA.  RNA purified using TRIzol reagent (Invitrogen) or the methods described in Preparation of RNA Using Guanidinium Isothiocyanate/Cesium Chloride Ultracentrifugation, Preparation of RNA from Paraffin-Embedded Fixed Tissue. - [Read Real-Time RT-PCR: cDNA Synthesis Protocol]

Category: PCR Protocols: Reverse Transcriptase RT-PCR Protocols

Protocol uses the Superscript II First-Strand Synthesis system for the generation of cDNA from total RNA.  RNA purified using TRIzol reagent (Invitrogen) or the methods described in Preparation of RNA Using Guanidinium Isothiocyanate/Cesium Chloride Ultracentrifugation, Preparation of RNA from Paraffin-Embedded Fixed Tissue. - [Read Real-Time RT-PCR: cDNA Synthesis Protocol]