Destaining electrophoretic

Detection and analysis Once the electrophoretic separation is completed, the gel is treated with a stain, most popularly Coomassie Brilliant Blue, to reveal protein bands. In the case of Coomassie Brilliant Blue, this process involves a stain-destain cycle, and as proteins can differ in their affinity to take up this dye the detection is really only considered qualitative. There are various other alternative stains which include Amido black. Ponceau red and silver nitrate. Of these, silver staining is widely considered to be one of the most sensitive staining... 

Fig. 7.15. Electrophoretic destaining apparatus (Richards and Gratzer 1968) cross sectional diagram of electrolytic destainer. Three gels are shown in the gel compartment (b) composed of a perspex (Incite) frame bounded by two sheets of Vyon porous plastic (d). This compartment rests in a holder made of two further perspex frames (a) joined along their bottom and sides. The gel compartment and the porous plastic are held in the holder by a fourth removable perspex frame (c). The holder fits tightly into a perspex tank (e) and thus separates the two buffer compartments (g), which are normally filled with 10% acetic acid. The potential is applied through two j in carbon rods (f) which are mounted in the tank through rubber grommets.

Several types of apparatus for electrophoretic destaining have been devised, and two designs have been shown in Figs. 7.15 and 7.16. In... 

The photopolymerization of the spacer gel is effected, the comb is carefully removed (in order not to destroy the surface of the wells), the plates are mounted into the electrophoretic apparatus, and samples are placed in the wells. After having filled the electrode vessels with buffer the electrophoretic separation can be started. Diffusion staining and destaining of the gel is done after disassembling the plates. 

It has been mentioned already that destaining is positively the most time-consuming operation. Frequent exchanging of the destaining solution is necessary to ensure obtaining the results within a reasonable period of time. In a most primitive way electrophoretic destaining can be materialized in a beaker filled with destaining solution into which both electrodes and the gel to be destained are placed. The efficiency of this system is, however, low in comparison with the more sophisticated devices either commercially available or described in the literature [215-222]. 

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