Specimen fragments are separated using a process called electrophoresis. This process takes advantage of the fact that DNA is highly negatively charged. If DNA molecules are placed in a gel matrix and an electric current is applied to the matrix, the DNA molecules will migrate to the positive electrode. To do this, a gel is first made between two pieces of optically pure glass. The gel is made from a chemical called acrylamide. This acrylamide is poured between the two pieces of glass that are held apart by 20 micrometer plastic spacers along their sides. Along the top of the gel another plastic spacer is used to create a straight flat edge where specimens will eventually be loaded. A catalyst is added to the liquid acrylamide mixture that causes it to polymerize into polyacrylamide. This polyacrylamide is a stiff clear gelatinous material that acts as a molecular sieve to separate DNA molecules in the process of electrophoresis. The apparatus used for this electrophoresis process has buffer chambers that attach to the top and the bottom of the gel between the two glass plates. One of the pieces of glass possesses a notch in which a plastic sharks tooth comb is inserted for loading of specimens. As one might expect, this sharks tooth comb has very sharp teeth that are inserted into the gel at the top of the gel apparatus. The specimens are then loaded in the spaces between the teeth.

Once the specimens are loaded, a very high voltage is applied to platinum electrodes in the buffer chambers. The upper chamber contains the negative electrode while the bottom chamber contains the positive electrode. Thus, the negatively charged DNA fragments are pushed through the gel by the current from the top to the bottom.

As the fluorescently labeled DNA fragments pass the laser excitation source near the bottom of the gel, they emit fluorescence at known wavelengths and are detected by a cooled CCD camera. This camera then downloads information to software that assembles a precise electronic representation of the gel in terms of intensity of fluorescent emission. This electronic image is processed using software that automatically finds specimen lanes and labels them. These data are then fed to a software analysis program which actually labels peaks of fluorescent emission as DNA fragments of known repeat size for each STR region.