When scientists need to separate and sort molecules in a sample according to their electric charge and size, they use a technique called gel electrophoresis. Gel electrophoresis is usually used to separate bio-molecules in DNA, RNA and proteins. The apparatus is essentially a box of gel with electrodes attached. The electrodes introduce an electric field to the gel that affect any samples placed in the gel. Specifically, positively-charged molecules are pulled toward the negative electrode (cathode) and negatively-charged molecules are pulled toward the positive electrode (anode). The amount of movement is logarithmically related to the size of each molecule – large molecules move more slowly than small ones. These different movement speeds create bands of materials in the gel, a sort of smearing out of the original sample into its constituent parts.
Not only does the gel physically support the separated samples, it disperses any heat buildup due to the electric current. Think of it as a gooey sieve, where large molecules have more trouble moving than do small ones. One form of the gel is extracted from seaweed and is called agar-agar. The sugar from agar-agar, called agarose, serves as a gel medium. While gels made of agar are used extensively, more sophisticated gels composed of cross-linked polymers are selected to match the expected molecular weights of the sample contents. For instance, small molecules like proteins and nucleic acids use a gel based on acrylamide together with a cross-linker, creating polyacrylamide. On the other hand, agarose is not cross-linked, but rather straight-chained, and is electrically neutral. It has large pores that allow larger molecules to pass.
Once materials have been separated in the gel, scientists often apply a stain to make them visible. Common stains include ethidium bromide (EtBr), silver and Coomassie Brilliant Blue (CBB). EtBr is used as a fluorescent stain for nucleic acids (DNA and RNA). If you shine ultraviolet light on EtBr, it will give off an orange glow that is magnified by the presence of DNA. Scientists protect their eyes when viewing samples under UV light by computer-recording the images as photographs. CBB is a triphenylmethane dye originally developed for coloring textiles. It is commonly used to visualize proteins. Originally, they stained the gel and the sample, and then de-stained the gel with vinegar after separation had occurred. Later on, they found they could mix the CBB with trichloracetic acid and use it on the sample only, no longer needing to stain and de-stain the gel.
The gel is often loaded with several parallel samples that form lanes of banding. Materials of the same size will migrate the same distance, so identical lanes indicate the material in each is the same or very similar. Scientists have developed molecular weight markers – molecules of known size – which serve as comparison bands when used in conjunction with samples of unknown size. Besides scientific research, gel electrophoresis is used by forensic investigators to identify evidence taken from crime scenes. Modern software programs are now used to help identify samples.