different methods to analyze samples for bacteria. Analysis Bacteria reproduce rapidly if conditions are right for growth. Most bacteria grow best in dark, warm, moist environments with food. When grown on solid media, some bacteria form colonies as they multiply which may grow large enough to be seen. By growing and counting colonies of fecal coliform bacteria from a sample of water, the amount of bacteria originally present can be determined. Membrane filtration is the method of choice for the analysis of fecal coliforms in water. Samples to be tested are passed through a filter of particular pore size (generally 0.45 micrometre). The microorganisms present in the water remain on the filter surface. The filter is placed in a sterile Petri dish with a selective medium, growth of the desired organisms is encouraged, while other non-target organisms is suppressed. Each cell develops into a separate colony, which can be counted directly, and the initial inoculum size can be determined. Typically sample volumes of 100 ml will be used for water testing and filtered, with the goal of achieving a final desirable colony density range of 20 to 60 colonies per filter. Contaminated sources may require dilution to achieve a "countable" membrane. The filter is placed on a Petri dish containing M-FC agar and incubated for 24 hours at 44.5 °C (112.1 degrees F). This elevated temperature heat shocks non-fecal bacteria and suppresses their growth. As the fecal coliform colonies grow they produce an acid (through fermenting lactose) that reacts with the aniline dye in the agar thus giving the colonies their blue color. Newer methods for coliform detection are based on specific enzyme substrates as indicators of coliforms. These assays make use of a sugar linked to a dye which, when acted on by the enzyme beta-galactosidase, produces a characteristic color. The enzyme beta-galactosidase is a marker for coliforms generally and may be assayed by hydrolysis of enzyme specific glycosides such as o-nitrophenyl-beta-D-galactose. Assays typically include a second sugar linked to a different dye which, when acted on by the enzyme beta-glucuronidase, produces a fluorescent product. Because E. coli produces both beta-galactosidase and beta-glucuronidase, a combination of two dyes makes it possible to differentiate and quantify coliforms and E. coli in the same pot. More recently, the chemistry behind enzymatic detection compounds has been updated so that the indicating component is redox active, as opposed to the more usual chromogenic