One-dimensional gels

Prokaryotic cells express hundreds to thousands of proteins while higher eukaryotes express thousands to tens of thousands of proteins at any given time. If these proteins are to be individually identified and characterized, they must be efficiently fractionated. One-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) has typically been use to study protein mixtures of <100 proteins. Onedimensional electrophoresis is useful because nearly all proteins are soluble in SDS, molecules ranging from approximately 10,000 to 300,000 molecular weight can be resolved, and extremely basic or acidic proteins can be visualized. The major disadvantage to one-dimensional gels is that they are not suitable for complex mixtures such as proteins from whole cell lysates. 

J Singh, ME Weber. Kinetics of one-dimensional gel swelling and collapse for large volume change. Chem Eng Sci 51 4499-4508, 1996. 

Optimal dilution of the primary and secondary antibody should be determined by immunoblotting of one-dimensional gels. Dot blot analysis can also be used. Working solutions of antibodies can be stored at -20°C and used several times (82). 

Fig. 1. Matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF) spectrum of a trypsin-digested one-dimensional gel band. Peaks are labeled with their monoisotopic masses. Note that these are not the masses of the peptides, but of the peptide (pseudo)molecular ions. In MALDI spectra, peptide molecular ions arise predominantly through the addition of a proton to the peptide, giving a mass increase of 1.007 Da. The molecular ions are usually denoted as MH+ or [M+H]+.

Camden, Australia An improved one-dimensional gel electrophoresis method was used to separate three waxy proteins in wheat starch 261... 

The arrangement used for two-dimensional gel electrophoresis may comprise various combinations of apparatus used for one-dimensional gel electrophoresis already described. Cylindrical gels used for the first dimension may be conveniently adapted to Akroyd type gels ( 8.4.2.1) (Martini and Gould 1971). Alternatively, strips of Akroyd... 

Figure 3. Example of Western blots of R and S Eleusine extracts probed with anti-sea urchin tubulin that recognizes both ot-and /1-tubulins. The R biotype has two 0-tubulin bands, but only one a-tubulin in one dimensional gel extracts.

In complex reactions on resins, one-dimensional gel-phase HR-MAS NMR spectroscopy does not always afford conclusive information to be obtained on the characterization of all the intermediates and final products. Two-dimensional HR-MAS NMR spectroscopy (COSY,TOCSY, HETCOR, and SECSY) provides crucial information on proton connectivity through / coupling information. With 2D HR-MAS NMR complete assignments can be made of spectra of complex bound-molecules, such as oligosaccharides and peptides, and even of noncovalent interactions between compounds connected to resins and receptor molecules. In sum, HR-MAS NMR spectroscopy has been successfully used for characterizing complex resin-bound products and intermediates. It has been used for analyzing biochemical compounds as well. Future work will adapt this technique to reaction monitoring and to routine quantitation besides its use as a characterization tool. 

Interaction of chemical waves with an applied electric field is an important feature of electrochemical systems [74], The influence of an external electric field on the precipitation of yellow and red mercuric iodide in one-dimensional gel media was studied by Das et al. [71] using an experimental setup, shown in Fig. 10.5a. In the absence of an electric field, the colour of the precipitate remained unchanged. In the presence of electric field, transitions from yellow to red (e, g) and red to yeUow (f, h) Hglj were obtained at the field intensity 0.567 Vcm. ... 

Treatment of either a tissue culture or a cell extract with Bodipy-epoxomicrn MVB-003 followed by SDS-PAGE readily reveals proteasome active sites (Figure 12.3c) [9, 10]. In case the constitutive proteasome is expressed exclusively, the three catalytic species pi, P2, and pS are readUy resolved. In case the treated tissue also expresses immunoproteasomes, the one-dimensional gel wUl resolve pi and p2i, but only partially pi, pii, pS, and pSi (see for a detailed experimental protocol Box 12.1 [11]). Two-dimensional gel electrophoresis, by which proteins are separated on the basis of the charge followed by mass allows for complete resolution of all six catalytic activities. Figure 12.3d represents an example of competitive ABPP. In this experiment, tissue or tissue extract is treated first with a prospective inhibitor. Ensuing incubation with... 

Figure 8 Canine serum. One-dimensional gel electrophoresis of dog serum separated on SDS-PAGE followed by silver staining. Track N is from a normai heaithy dog and track M is from a dog with a monocionai gammopathy (igA) Prominent H and L chains can be cieariy seen. (Courtesy of Miiier i, University of Veterinary Medicine, Vienna.)...

FIGURE 3 One-dimensional gel electrophoresis (Laemmli s, 15% acrylamide) of histone... 

Figure 3 shows one-dimensional gels of histones (Figs. 3A and B) and nonhistone proteins (Fig. 3C) run in a 15% acrylamide Laemmli gel and stained with Coomassie brilliant blue. Representative silver-stained gels of 16.5% T, 6% of proteins from three different cell types are shown in Fig. 4. Serva blue G migrates as a broad band in front of the smallest proteins in the gels as described here. This is not always the case for the commonly used bromophenol blue.

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