![]() ![]() “Normalize signal intensity to total protein loading (assessed by staining membranes for total protein) whenever possible” – Journal of Biological Chemistry.“Record how data were obtained, whether signal intensity was linear with antigen loading, and how protein loading was normalized” – Journal of Biological Chemistry.“For quantitative comparisons, appropriate reagents, controls and imaging methods with linear signal ranges should be used” – Nature.Many leading journals have developed guidelines for submitting western blotting research and select quotes from those guidelines are highlighted below. In recent years, journal editors and reviewers have been requesting adoption of methods that improve the accuracy and reproducibility of quantitative western blot data. Methods for protein normalization include use of internal housekeeping protein controls such as GAPDH, β-tubulin, β-actin, or cyclophilin B, exogenous loading controls, or total protein normalization. Protein normalization is used to correct for this variability and is a critical step in obtaining reliable and reproducible quantitative western blot data. In western blotting, variable results can be caused by unequal protein sample concentration, inconsistent sample loading onto the gel, and/or transfer variation during electroblotting. No-Stain Labeling Buffer, 20X (2 x 20 mL each storage 15-30⁰C) No-Stain Derivatizer (800 µL storage -20⁰C) No-Stain Activator (800 µL storage -20⁰C) The No-Stain Protein Labeling Reagent forms covalent bonds with a portion of the lysine amino acid side chains on all protein samples Tunable sensitivity-Increasing incubation time or doubling the reagent concentration produces a proportionally stronger signal for improved detection of low abundance proteins.Ī standard kit will label 40 mini gel-sized membranes or 40 mini gels, or a combination of the twoĬompatible with any gel type (precast or pour your own) or gel chemistry (Bis-Tris, Tris-glycine, Tris-acetate, Tricine)Ĭompatible with all downstream immunodetection steps with chemiluminescence or fluorescence-based detectionĬompatible with a wide range of imagers that have UV or fluorescence light sources, for example, the iBright FL1500 Imaging System.Sensitive and stable signal-Protein bands are detected down to 20 ng and the signal is compatible with downstream antibody detection.Broad linear dynamic range-1–80 µg total protein loaded per well. ![]()
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