recommended Protocols

Category: Reporter Gene Assays: B-Galactosidase Assay Protocols

The recommended amount of RSV-ß-Galactosidase plasmid to use for transfection of cells (60 mm or 100 mm dish) is 1-2 µg. The optimal amount of plasmid DNA will be determined by the efficiency of transfection , which is very dependent upon the particular cell line and transfection protocol. - [Read B- Galactosidase Assay Protocol]

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]

Category: Cell Culture Protocols: Cell Fixing Protocols

Treating cells with paraformaldehyde leads to the establishment of chemical cross-links between free amino groups. When the cross-links join different molecules, a latticework of interactions occurs that holds the overall architecture of the cell together. Commercial formaldehyde solutions are not recommended, because they lack the advantages of using a variable-length polymer, and the cells will simultaneously be fixed with the alcohol (usually methanol). - [Read Fixing Attached Cells in Paraformaldehyde Protocol]

Category: Chromatography Protocols: Affinity Chromatography Protocols

Two methods are provided for purifying glycoproteins using wheat-germ agglutinin or concanavalin A-Sepharose.  Because lectin-affinity matrices can bind a few milligrams of protein per milliliter of affinity matrix, only a small amount of affinity gel matrix is required.  The batchwise method is recommended when protein volume is large. - [Read Lectin-Agarose Affinity Chromatography Protocol]

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from large-scale (500 ml) bacterial cultures by treatment with Triton X-100 and lysozyme, followed by heating.  This method is not recommended for preparing plasmid DNA from strains of E. coli that express endonuclease A (endA+ strains). - [Read Preparation of Plasmid DNA by Large-scale Boiling Lysis Protocol]

Category: Nucleic Acid Purification Protocols: DNA Isolation Protocols: DNA Plasmid Preps Protocols

Plasmid DNA is isolated from small-scale (1-2 ml) bacterial cultures by treatment with Triton X-100 and lysozyme, followed by heating.  This method is not recommended for preparing plasmid DNA from strains of E. coli that express endonuclease A (endA+ strains). - [Read Preparation of Plasmid DNA by Small-scale Boiling Lysis Protocol]

Category: Chromatography Protocols: DNA RNA Affinity Chromatography

Chromatography on oligo(dT) columns is the preferred method for large-scale purification (>25 µg) of poly(A)+ RNA extracted from mammalian cells. Typically, between 1% and 10% of the RNA applied to the oligo(dT) column is recovered as poly(A)+ RNA. Because the method can be frustratingly slow, it is not recommended for purification of poly(A)+ RNA from multiple samples. For this purpose, batch elution (Selection of Poly(A)+ RNA by Batch Chromatography) is the better choice. - [Read Selection of Poly(A)+ RNA by Oligo(dT)-Cellulose Chromatography - Subscription Required]