ArticleBMC immunology2014
Rapid preparation of high-purity nuclear proteins from a small number of cultured cells for use in electrophoretic mobility shift assays.
Article in BMC immunology, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 16 papers, 1 of them a synthesis that pooled it.
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Who cites it
16 citing papers in PubMed, 1 synthesis or guideline pooled it, 26 citations in OpenAlex.
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- Toward G-Quadruplex-Based Anticancer Agents: Biophysical and Biological Studies of Novel AS1411 Derivatives.International journal of molecular sciences · 2020Article
- Methylglyoxal-derived advanced glycation end products induce matrix metalloproteinases through activation of ERK/JNK/NF-κB pathway in kidney proximal epithelial cells.Food science and biotechnology · 2020Article
- Persistent pulmonary hypertension alters the epigenetic characteristics of endothelial nitric oxide synthase gene in pulmonary artery endothelial cells in a fetal lamb model.Physiological genomics · 2018Article
- Versatile and efficient chromatin pull-down methodology based on DNA triple helix formation.Scientific reports · 2018Article
- Measurement of NF-κB Activation in TLR-Activated Macrophages.Methods in molecular biology (Clifton, N.J.) · 2018Article
- Conditional, Genetically Encoded, Small Molecule-Regulated Inhibition of NFκB Signaling in RPE Cells.Investigative ophthalmology & visual science · 2017Article
Corrections and comments
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Authors and funding
9 authors at 2 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundHighly purified nuclear protein is required when using an electrophoretic mobility shift assay (EMSA) to study transcription factors, e.g. nuclear factor-κB (NF-κB), a major transcription factor that regulates both innate and adaptive immune responses following infection. Although many protocols have been developed for nuclear protein extraction, they are not necessarily optimized for use in EMSA, often require a large number of cells and long processing times, and do not always result in complete separation of the nuclear and cytoplasmic fractions.
resultsWe have developed a simple, rapid and cost-effective method to prepare highly purified nuclear proteins from a small number of both suspended and adherent cultured cells that yields nuclear proteins comparable to those prepared by a standard large-scale method. The efficiency of the method was demonstrated by using EMSA to show the successful detection, in multilple concurrent samples, of NF-κB activation upon tetradecanoyl phorbol acetate (TPA) stimulation.
conclusionsThis method requires only a small number of cells and no specialized equipment. The steps have been simplified, resulting in a short processing time, which allows researchers to process multiple samples simultaneously and quickly. This method is especially optimized for use in EMSA, and may be useful for other applications that include proteomic analysis.
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