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Gel electrophoresis
About: Gel electrophoresis is a research topic. Over the lifetime, 26026 publications have been published within this topic receiving 1113565 citations.
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TL;DR: These proteins were released from labile cortical regions of animal cells in response to perturbations of homeostasis in cells as evolutionarily distinct as cultured rat embryo cells and squid glial cells, supporting the conclusion that a selective release mechanism is involved.
Abstract: Cultured rat embryo cells were stimulated to rapidly release a small group of proteins that included several heat-shock proteins (hsp110, hsp71, hscp73) and nonmuscle actin. The extracellular proteins were analyzed by two-dimensional polyacrylamide gel electrophoresis. Heat-shocked cells released the same set of proteins as control cells with the addition of the stress-inducible hsp110 and hsp71. Release of these proteins was not blocked by either monensin or colchicine, inhibitors of the common secretory pathway. A small amount of the glucose-regulated protein grp78 was externalized by this pathway. The extracellular accumulation of these proteins was inhibited after they were synthesized in the presence of the lysine analogue aminoethyl cysteine. It is likely that the analogue-substituted proteins were misfolded and could not be released from cells, supporting our conclusion that a selective release mechanism is involved. Remarkably, actin and the squid heat-shock proteins homologous to rat hsp71 and hsp110 are also among a select group of proteins transferred from glial cells to the squid giant axon, where they have been implicated in neuronal stress responses (Tytell et al.: Brain Res., 363:161-164, 1986). Based in part on the similarities between these two sets of proteins, we hypothesized that these proteins were released from labile cortical regions of animal cells in response to perturbations of homeostasis in cells as evolutionarily distinct as cultured rat embryo cells and squid glial cells.
389 citations
TL;DR: The comet assay has been modified to detect various base alterations, by including digestion of nucleoids with a lesion-specific endonuclease, and modifications to measure cellular antioxidant status and different types of DNA repair.
Abstract: The comet assay (single cell gel electrophoresis) is the most common method for measuring DNA damage in eukaryotic cells or disaggregated tissues. The assay depends on the relaxation of supercoiled DNA in agarose-embedded nucleoids (the residual bodies remaining after lysis of cells with detergent and high salt), which allows the DNA to be drawn out towards the anode under electrophoresis, forming comet-like images as seen under fluorescence microscopy. The relative amount of DNA in the comet tail indicates DNA break frequency. The assay has been modified to detect various base alterations, by including digestion of nucleoids with a lesion-specific endonuclease. We describe here recent technical developments, theoretical aspects, limitations as well as advantages of the assay, and modifications to measure cellular antioxidant status and different types of DNA repair. We briefly describe the applications of this method in genotoxicity testing, human biomonitoring, and ecogenotoxicology.
389 citations
TL;DR: The amino acid composition of protein G was determined and was found to be different from that of protein A, the well known staphylococcal IgG-binding protein, and the binding between protein G and various polyclonal and monoclonal IgG was pH dependent and strongest at pH 4 and 5, and weakest at pH 10.
388 citations
TL;DR: With this method 1 ng of active enzyme can easily be detected and both prokaryotic and eukaryotic cell extracts can be examined, and changes in the size of enzymatically active proteins can be determined.
387 citations
TL;DR: The ratios of the oxidative phosphorylation complexes NADH:ubiquinone reductase (complex I), succinate:UBiquin one reduct enzyme (complex II), ubiquinol:cytochrome c reductases (complex III), cy tochrome c oxidase ( complexes IV), and F1F0-ATP synthase ( complex V) from bovine heart mitochondria were determined by applying three novel and independent approaches that gave consistent results.
385 citations