Molecular weight determinations and the influence of gel density, protein charges and protein shape in polyacrylamide gel electrophoresis
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Polyacrylamide
Molecular mass
Bovine serum albumin
Molecular-weight size marker
PurposeTo establish a modified electrophoresis method for the analysis of peptides with low molecular weights in Atranticance.MethodsInvestigating the effects of different factors. The linear regression analysis was made using molecular markers and the molecular weights of the components in Atranticance were determined.ResultsA method was established. The relative molecular weights of the components in the sample are 23 500,15 400,13 400,11 300,9 300,8 000 respectively.ConclusionSeparating gel with 6 mol/L urea at higher acrylamide concentration (16.5% T,6% C) is useful for identification of middle and low molecular peptides and is a well-established method for the analysis of middle and low molecular peptides.
Molecular mass
Sodium dodecyl sulfate
Molecular-weight size marker
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A simple method for detection of DNA-binding proteins is offered. These proteins can be revealed, following their electrophoretic separation in sodium dodecyl sulfate (SDS)-polyacrylamide gel containing labeled DNA, by washing the gel in buffer to remove SDS and to allow protein renaturation. Protein-free DNA is washed out, remaining in the DNA-binding proteins that restored their original characteristic. After autoradiography these proteins are seen as black bands (by one-dimensional gel electrophoresis) or spots (by two-dimensional gel electrophoresis) on a grey background. High sensitivity of the method is shown by using protein fractions of rat liver and a standard method.
Molecular-weight size marker
Sodium dodecyl sulfate
Free-flow electrophoresis
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Abstract Electrophoresis is used to separate complex mixtures of proteins (e.g., from cells, subcellular fractions, column fractions, or immunoprecipitates), investigate subunit compositions, verify homogeneity of protein samples, and purify proteins for use in further applications. In polyacrylamide gel electrophoresis, proteins migrate in response to an electrical field through pores in a polyacrylamide gel matrix; pore size decreases with increasing acrylamide concentration. The combination of pore size and protein charge, size, and shape determines the migration rate of the protein. In this unit, the standard Laemmli method is described for discontinuous gel electrophoresis under denaturing conditions, i.e., in the presence of sodium dodecyl sulfate (SDS). Support protocols cover the casting of gels, calculation of molecular mass using the electrophoretic mobility of a protein, and purification of SDS by recrystallization.
Molecular-weight size marker
Sodium dodecyl sulfate
Electrochromatography
Free-flow electrophoresis
Protein purification
Polyacrylamide
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Color marker
Molecular-weight size marker
Polyacrylamide
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Apolipoproteins B100 and B48 in human and rat plasma were studied by using sodium dodecyl sulfate (SDS) polyacrylamide gradient gel electrophoresis. On SDS gradient gel electrophoresis, human and rat apoprotein B100 co-migrated and had an apparent Mr=258,000±12,000. Human and rat apoprotein B48 had an apparent Mr=189,000±6,000. The molecular weight of human apoprotein B100 determined by sedimentation equilibrium analysis was 270,000±20,000, which was similar to the value determined by SDS gradient gel electrophoresis. However, on SDS polyacrylamide gel electrophoresis at constant concentration, the relative migration value of human apoprotein B100 was not constant when the concentration of polyacrylamide was changed. These results indicate that SDS gradient gel electrophoresis is more suitable for the analysis of apolipoprotein B's than ordinary SDS polyacrylamide gel electrophoresis.
Sodium dodecyl sulfate
Molecular-weight size marker
Polyacrylamide
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Molecular mass
Molecular-weight size marker
Sodium dodecyl sulfate
Ribosomal protein
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ABSTRACT The most widely used Western blotting protein standards are prestained proteins of known molecular mass (kDa). They are also utilized for sodium dodecyl sulphate (SDS) Polyacrylamide Gel Electrophoresis (PAGE) to determine the molecular mass of proteins separated by electrophoresis. The objective of this study was to assess the reliability of different commercially available protein standards in predicting accurate protein molecular weights. We performed this experiment by running Criterion TGX gels with five prestained protein standards (Thermo Fisher SeeBlue Plus 2, Bio-Rad Precision Plus Protein Dual-color, Thermo Fisher Spectra Multi-color, Novex-Sharp Pre-stained, and Invitrogen iBright Pre-Stained). To evaluate their accuracy, we utilized highly purified Bovine Serum Albumin (BSA, 66.44 kDa) and Cytochrome C (Cyto C, 11.62 kDa). We also made use of the dimers of BSA (132.88 kDa) and Cyt C (23.24 kDa) that are present on SDS-PAGE gels. Our results suggest that three of the standards were less accurate at higher molecular masses with the iBright marker having the highest error in determining the expected 132.88 kDa molecular weight. The SeeBlue Plus 2 was accurate at identifying the 132.88 kDa molecular weight protein band but was less reliable for the three other lower molecular weight proteins. These findings have significant implications for the determination of protein masses because researchers rely on these standards to evaluate the molecular masses of their protein(s). We suggest that at least two different protein standards should be initially used in electrophoresis gels and for Western blotting in order to get accurate protein molecular weight results.
Molecular mass
Molecular-weight size marker
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Molecular-weight size marker
Sodium dodecyl sulfate
Polyacrylamide
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Abstract A miniature system for two‐dimensional polyacrylamide gel electrophoresis is presented. Discontinuous sodium dodecyl sulfate gel electrophoresis in micro‐slab gels following high resolution micro‐electrophoresis in cylindrical continuous polyacrylamide gradient gels separates macromolecules after an initial size fractionation into their subunits. The construction of a glass cuvette for micro‐slab gels and of a suitable electrophoresis tank is described in detail. Furthermore, procedures for silver staining and fluorography initially developed for gels of normal size were adapted to micro‐slab gels. Crude tissue extracts of Funaria hygrometrica L. Sibth. were analyzed without any further purfication. Twenty samples can be processed in about eight hours.
Molecular-weight size marker
Polyacrylamide
Sodium dodecyl sulfate
Free-flow electrophoresis
Cuvette
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