Abstract
Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and nonspecific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein recovery efficiency. Here, we describe the enhancement of protein separation efficiency for up to twofold in conventional one-dimensional PAG electrophoresis (1D PAGE), two-dimensional (2D) PAGE, and native PAGE by implementing pulses of inverted electric field during gel electrophoresis.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
Schwartz DC, Cantor CR (1984) Separation of yeast chromosome sized DNA fragments by pulsed field gradient gel electrophoresis. Cell 37:67–75
Bancroft I, Wolk CP (1988) Pulsed homogeneous orthogonal field gel electrophoresis (PHOGE). Nucleic Acids Res 16:7405–7418
Carle GF, Frank M, Olson MV (1986) Electrophoretic separations of large DNA molecules by periodic inversion of the electric field. Science 232:65–68
Chu G, Vollrath D, Davis RW (1986) Separation of large DNA molecules by contour-clamped homogeneous electric fields. Science 234:1582–1585
Bostock CJ (1988) Parameters of field inversion gel electrophoresis for the analysis of pox virus genomes. Nucleic Acids Res 16:4239–4252
Heller C, Pohl FM (1989) A systematic study of field inversion gel electrophoresis. Nucleic Acids Res 17:5989–6003
Mathew MK, Hui CF, Smith CL et al (1988) High-resolution separation and accurate size determination in pulsed-field gel electrophoresis of DNA. 4. Influence of DNA topology. Biochemistry 27:9222–9226
Mathew MK, Smith CL, Cantor CR (1988) High-resolution separation and accurate size determination in pulsed-field gel electrophoresis of DNA. 2. Effect of pulse time and electric field strength and implications for models of the separation process. Biochemistry 27:9210–9216
Mathew MK, Smith CL, Cantor CR (1988) High-resolution separation and accurate size determination in pulsed-field gel electrophoresis of DNA. 1. DNA size standards and the effect of agarose and temperature. Biochemistry 27:9204–9210
Tischfield JA, Bernhard HP, Ruddle FH (1973) A new electrophoretic-autoradiographic method for the visual detection of phosphotransferases. Anal Biochem 53:545–554
Brassard E, Turmel C, Noolandi J (1991) Observation of orientation and relaxation of protein-sodium dodecyl sulfate complexes during pulsed intermittent field polyacrylamide gel electrophoresis. Electrophoresis 12:373–375
Houri A, Starita-Geribaldi M (1994) Pulsed field electrophoresis for the separation of protein-sodium dodecyl sulfate-complexes in polyacrylamide gels. Electrophoresis 15:1032–1039
Sant'Ana Pereira JA, Greaser M, Moss RL (2001) Pulse electrophoresis of muscle myosin heavy chains in sodium dodecyl sulfate-polyacrylamide gels. Anal Biochem 291:229–236
Rodbard D, Chrambach A (1970) Unified theory for gel electrophoresis and gel filtration. Proc Natl Acad Sci U S A 65:970–977
Guo XH, Chen SH (1990) Reptation mechanism in protein-sodium-dodecylsulfate (SDS) polyacrylamide-gel electrophoresis. Phys Rev Lett 64:2579–2582
Kozulic B (1994) On the “door-corridor” model of gel electrophoresis. I. Equations describing the relationship between mobility and size of DNA fragments and protein-SDS complexes. Appl Theor Electrophor 4:125–136
Tsai H, Low TY, Freeby S et al (2007) Increase in local protein concentration by field-inversion gel electrophoresis. Proteome Sci 5:18
Acknowledgment
This work was supported by the Agency for Science Technology and Research (ASTAR) of Singapore. BioMed Central is the original publisher of the original research article [18]. Figures are used with permission of BioMed Central.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Tsai, H., Leung, HC.E. (2019). Increase in Local Protein Concentration by Field-Inversion Gel Electrophoresis. In: Kurien, B., Scofield, R. (eds) Electrophoretic Separation of Proteins. Methods in Molecular Biology, vol 1855. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8793-1_19
Download citation
DOI: https://doi.org/10.1007/978-1-4939-8793-1_19
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-8792-4
Online ISBN: 978-1-4939-8793-1
eBook Packages: Springer Protocols