Two dimensional-polyacrylamide gel electrophoresis

Fawcett, JS Sullivan, SV Chrambach, A, Toward a Steady-State Pore Limit Electrophoresis Dimension for Native Proteins in Two-Dimensional Polyacrylamide Gel Electrophoresis, Electrophoresis 10, 182, 1989. 

Pietrogrande, M.C., Marchetti, N., Tosi, A., Dondi, E, Righetti, P.G. (2005). Decoding two-dimensional polyacrylamide gel electrophoresis complex maps by autocovariance function a simplified approach for proteomics. Electrophoresis 26, 2739-2748. 

Scharfe C et al. MITOP, the mitochondrial proteome database 2000 update. Nucleic Acids Res 2000 28 155-158. Molloy MP et al. Establishment of the human reflex tear two-dimensional polyacrylamide gel electrophoresis reference map new proteins of potential diagnostic value. Electrophoresis 1997 18 2811-2815. 

Figure 7.1. Effect of GM-CSF on neutrophil protein biosynthesis. Human neutrophils were incubated in RPMI1640 medium supplemented with 2.5% foetal calf serum and 60 jtCi/ml [35S]-methionine, in the presence and absence of 50 U/ml GM-CSF. After 4 h incubation at 37 °C, the cells were pelleted by low-speed centrifugation. The proteins in the cell pellets were precipitated by 10% trichloracetic acid and then analysed by two-dimensional polyacrylamide gel electrophoresis, using iso-electrofocussing in the first dimension. Electrophoresis in the second dimension was performed in the presence of SDS and used a 12% acrylamide gel. Source Experiment of Becky Stringer.

Orphanides G, LeRoy G, Chang CH, Luse DS, Reinberg D (1998) FACT, a factor that facilitates transcript elongation through nucleosomes. Cell 92 105-116 Orrick LR, Olson MO, Busch H (1973) Comparison of nucleolar proteins of normal rat Uver and Novikoff hepatoma ascites cells by two-dimensional polyacrylamide gel electrophoresis. Proc Natl Acad Sci USA 70 1316-1320... 

Two-Dimensional Polyacrylamide Gel Electrophoresis (2D-PAGE lEF followed by SDS-PAGE)... 

Craven BA, Totty N, Harnden P, Selby PJ, Banks RE. (2002) Laser capture microdissection and two-dimensional polyacrylamide gel electrophoresis evaluation of tissue preparation and sample limitations. Am J Pathol 160, 815-22. 

Dunn, M.J. Corbett, J.M. (1996) Two-dimensional polyacrylamide gel electrophoresis. Methods Enzymol. 271, 177-203. 

Proteomics Chromatographic Fractionation Prior to Two-Dimensional Polyacrylamide Gel Electrophoresis for Enrichment of Low-Abundance Proteins to Facilitate Identification by Mass Spectrometric Methods... 

NHPacker, A Pawlak, WC Kett, AA Gooley, JW Redmond, KL Williams. Proteome analysis of glycoforms a review of strategies for the microcharacterization of glycoproteins separated by two-dimensional polyacrylamide gel electrophoresis. Electrophoresis 18 452-460, 1997. 

LB Epstein, DM Smith, NM Matsui, HM Tran, C Sullivan, I Raineri, AL Burlingame, KR Clauser, SC Hall, LE Andrews. Identification of cytokine-regulated proteins in normal and malignant cells by the combination of two-dimensional polyacrylamide gel electrophoresis, mass spectrometry, Edman degradation and immunoblotting and approaches to the analysis of their functional roles. Electrophoresis 17 1655-1670, 1996. 

D Lutomski, M Caron, J-D Cornillot, P Bourin, C Dupuy, M Pontet, D Bladier, R Joubert-Caron. Identification of different galectins by immunoblotting after two-dimensional polyacrylamide gel electrophoresis with immobilized pH gradients. Electrophoresis 17 600-606, 1996. 

Before examining contemporary proteomic workflows, it must be said that the older gel-based technology still remains the largest contributor of proteomic-level information. For many years, the main means to analyze complex protein samples involves separation of proteins on the basis of size and isoelectric point (pi) using two-dimensional polyacrylamide gel electrophoresis (2-DE). The densitometric aspects of gel protein spots provide a direct means to visualize and quantify proteins based on the experimental conditions applied to the... 

Zannis and Breslow extended this analysis with two-dimensional polyacrylamide gel electrophoresis of apoE (isoelectric focusing in the first dimension followed by SDS-polyacrylamide gel electrophoresis in the second dimension). They numbered the isoprotein positions 1 to 7 from basic to... 

More protein sequences will be deduced from DNA sequences in the future. However, the sequencer utilizing the principle of Edman degradation wil continue to be an important tool. Sensitivity should be enhanced with further development of fluorescent reagents. As enormous improvement has made in recent years, mass spectrometry is quite promising for its role in sequence determination. Two-dimensional polyacrylamide gel electrophoresis and capillary electrophoresis show excellent resolution and detection sensitivity. 

Significant improvements in the technologies of high-resolution two-dimensional Polyacrylamide Gel Electrophoresis (2-D PAGE) and Mass Spectrometry (MS) have marked the start of proteome analysis. Proteomics permits the analysis of thousand of proteins simultaneously, and have the potential to identify markers for early detection, classification and prognosis of diseases, as well as pinpointing targets for improved treatment outcomes [42]. 

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