Electrophoresis apparatus

Stalcup aiid co-workers [14] adapted this method to a continuous elution mini-prep electrophoresis apparatus shown in Fig. 11-3. In this apparatus, the end of the electrophoretic gel is continuously washed with elution buffer. The eluent can then be monitored using an HPLC detector (Fig. 11-4) and sent to a fraction collector where the purified enantiomers, as well as the chiral additive, may be recovered. In this system, the gel configuration was approximately 100 mm x 7 mm, and was aircooled. The number of theoretical plates obtained for 0.5 mg of piperoxan with this gel was approximately 200. A larger, water-cooled gel was able to handle 15 mg of... 

Fig. 11-5. Schematic of continuous free flow electrophoresis apparatus.

As in CE, changing system variables (e.g., pH, ionic strength, additive concentration) is very easy in any of the continuous free flow electrophoresis systems reported here because all the interactions take place in free solution. Indeed, changing system variables may be easier in continuous free flow electrophoresis systems than in a CE system because there are essentially no wall effects. Of course, changing system variables in the continuous free flow electrophoresis apparatus may also be easier... 

Douglas et al. [98] have measured protein (serum albumin, ovalbumin, and hemoglobin) mobilities over a range of pH values using a free-flow electrophoresis apparatus and a particle electrophoresis apparatus. They found good agreement between the two measurements however, they also found some differences between their measurements and those reported in the older literature. They attributed the differences to the use of moving-boundary electrophoresis methods in the early experimental work and to differences in... 

Hjerten, S., Elenbring, K., Kilar, F., Liao, J.-L, Chen, A. J. C., Siebert, C. J., and Zhu, M.-D., Carrier-free zone electrophoresis, displacement electrophoresis and isoelectric focusing in a high-performance electrophoresis apparatus, /. Chromatogr., 403, 47, 1987. 

In most experiments the smallest amount of electrolyte needed to coagulate the sols measured after 2 hours standing was chosen as the CCC. When using HC1, this point is the critical coagulation pH. A constant temperature water bath was used for temperature different than 23°C. The pH values were measured with a Beckman Model 96A pH meter and a Fisher combination electrode. The electrophoretic mobility measurements were made with a Laser Doppler Electrophoresis apparatus. These experiments were performed by Mr. J. Klein of the Chemistry Department, Syracuse University. 

Another separation technique utilizes an electric field. An electric held is an electrically charged region of space, such as between a pair of electrodes connected to a power supply. The technique utilizes the varied rates and direction with which different organic ions (or large molecules with charged sites) migrate while under the influence of the electric held. This technique is called electrophoresis. Zone electrophoresis refers to the common case in which a medium such as cellulose or gel is used to contain the solution. A schematic diagram of the electrophoresis apparatus resembles an electrochemical apparatus in many... 

Figure 11.22 represents a paper electrophoresis apparatus. The soaked cellulose sheet is sandwiched between two horizontal glass plates with the ends dipped into vessels containing more electrolyte solution. The electrodes are also dipped into these vessels, as shown. The sample is spotted in the center of the sheet, and the oppositely charged ions then have room to migrate in opposite directions on the sheet. Qualitative analysis is performed much as with paper chromatography, by comparing the distances the... 

A typical gel electrophoresis apparatus is shown in Figure 11.23. The thin gel slab referred to above is contained between two glass plates. The slab is held in a vertical position and has notches at the top where the samples to be separated are spotted or streaked. In the configuration shown in the figure, only downward movement takes place, and thus only one type of ion, cation or anion, can be separated, since there is only one direction to go from the notch. 

This experiment requires a horizontal electrophoresis apparatus, power supply, and a gel-pouring tray with a comb to form sample wells. The FB 1001 apparatus available from Fisher Biotech and an apparatus available from EdVotek both are appropriate. The gel-pouring tray with combs is also available from EdVotek. Directions for preparing the tray accompany the unit. The Fisher apparatus includes a cooling unit that allows running at a constant temperature. In the EdVotek apparatus, the gel is under buffer. Since samples are loaded after the gel is immersed in buffer, sucrose must be added to increase sample density. 

Remove the comb carefully, and then remove the tape or sealing apparatus from both ends of the tray. Carefully let the gel slide out of the tray onto the electrophoresis apparatus. 

Add buffer to electrode chambers of the electrophoresis apparatus. Set up the electrodes so that the positive pole is opposite the loaded wells. Run the electrophoresis at 60 V for 40 min. 

Capillary procedures offer several advantages, including speed, resolution, sensitivity and technical simplicity, compared with the traditional methods on which they were based. An added advantage for iso-electric focusing in capillaries is the fact that it can be performed without a gel, but a coating on the internal surface of the capillary is usually required to reduce electroendosmosis. Similarly, isotachophoresis can be conveniently performed in capillary electrophoresis apparatus. 

L. R. Middendorf, J. C. Bruce, R. C. Bruce, R. D. Eckles, D. L. Grone, S. C. Roemer, G. D. Sloniker, D. L. Steffens, S. L. Sutter, J. A. Brumbaugh and G. Patonay, Continuous, on-line DNA sequencing using a versatile infrared laser scanner/electrophoresis apparatus Electrophoresis 13, 487-494... 

An imager is the most significant investment of all electrophoresis apparatus. As with all significant purchases, comparison shopping among the available products is highly recommended. In practice, researchers access the data in their... 

Wait for 5-lOmin and let the gel solidify. Then, remove the combs slowly and assemble plates containing the gel in the designed place in the vertical electrophoresis apparatus. 

Add 800-900mL IX TBE buffer into the space in the electrophoresis apparatus up to 5mm above the upper edge of the gel. 

Figure 3.7 contains an illustration of the basic components of a typical electrophoresis apparatus. The troughs at either end contain an electrolyte buffer solution. The sample to be separated is placed in the approximate center of the electrophoresis strip. 

The gel cassette is mounted into the electrophoresis apparatus and electrophoresis tanks are filled with electrode buffer (Soln. H). Remove air bubbles at the interfaces between electrode buffer and... 

Connect the electrophoresis apparatus to the power supply and switch on the voltage (time course) directly after sample application. 

Mount the gel into the electrophoresis apparatus fill the electrode chambers with 1 20 diluted MBS or MOPS electrode buffer. 

Put the plate on the cooling plate of a horizontal electrophoresis apparatus. Fill the buffer chambers with 1 1 diluted Soln. B (= electrode buffer) and connect the electrode buffer to the gel by filter paper bridges, wetted with electrode buffer. The filter paper covers the gel for about 5 mm. Remove carefully air bubbles between papers and gel. 

Apply some drops of kerosene on the cooling block of the horizontal electrophoresis apparatus and apply the IPG strips with the acidic side to the anode. Make 1 cm broad strips from 2 mm thick filter paper. Wet the strips with deionized water (no ultra-pure water ) and place them on top of the IPG strips as well as at the anodic (acidic) and the cathodic (basic) ends of the strip(s). 

Place the electrodes and press them gendy on the lEF electrode strips. Close the lid of the electrophoresis apparatus and switch on voltage. For 180 mm IPG strips the following conditions should be used (Table 2.11) ... 

The agarose (Soln. B) is molten in aboiling water bath and cooled to about 50 °C. Soln. D is added to a final concentration of 0.5 pg/mT . The liquid agarose is poured into the gel chamber of a horizontal electrophoresis apparatus and the comb is inserted. 

The connection between electrode chambers of a horizontal electrophoresis apparatus filled with buffer A and the cellulose support is made by wetted paper bridges. To avoid liquid moving, the buffer level must be the same in both chambers. A glass plate, laying on the paper bridges, covers the separation plate cooled to 0-4 °C. 

Fill the gel particles together with the surrounding buffer into the tube. The tube is inserted into the vertical electrophoresis apparatus (dialysis membrane down to anode), electrode buffer is poured, and electroelution is started with 10 mA per tube for 3-8 h, depending on tbe molar mass of the protein. 

Place the slide into a horizontal electrophoresis apparatus, fill the tanks with Soln. A, and connect the small ends of the slide to the electrode tanks by moistened filter paper wicks. Fill the well directed towards the anode with antiserum dilution and pipet antigen solution containing traces of bromophenol blue in that well which is nearby tbe cathode. 

FIGURE 3.2 A slab gel electrophoresis apparatus. A voltage source generates an electric potential difference between the upper and lower buffer chambers, causing the applied DNA sample to migrate through the gel toward the positive electrode. 

Rehydrated strips are then placed onto a custom-made cooling plate or a commercially available strip tray of a horizontal-type electrophoresis apparatus. The strip tray is fitted with bars holding the electrodes. The bars fitted with sample cup onto the tray allow easy application of the... 

FIG. 12.11 Schematic illustration of a Tiselius-type moving boundary electrophoresis apparatus. 

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