Endo Broth is used in the standard total Coliform membrane filtration procedure for testing water and bottled water. It is also used in the delayed-incubation total coliform procedure by adding sodium benzoate to make m Endo preservative medium (ref. 2). The coliform bacteria are defined as bacteria that produce a red colony with a metallic sheen within 24 hours incubation at 35°C on an Endo-type medium.
Appearance:Grayish purple to pinkish purple, homgenous, free flowing powder
Solubility (4.8%):Grayish purple, opalescent, with precipitate
pH (4.8%):7.2±0.1
Principles of the Procedure:Endo Broth contains peptones as sources of carbon, nitrogen, vitamins and minerals. Yeast extract supplies B-complex vitamins, which stimulate bacterial growth. Lactose is the carbohydrate. Phosphates are buffering agents. Sodium chloride maintains the osmotic balance of the medium. Sodium desoxycholate and sodium lauryl sulfate are added as inhibitors of gram-positive bacteria. Basic fuchsin is a pH indicator. Sodium sulfite is added to decolorize the basic fuchsin solution. The ethanol additive increases the antibacterial nature of the formulation
Lactose-fermenting bacteria produce acetaldehyde that reacts with the sodium sulfite and fuchsin to form red colonies. The development of a metallic sheen occurs when the organism produces aldehydes with the rapid fermentation of lactose. If the inoculum is too heavy, the sheen will be suppressed. Lactose-nonfermenting bacteria form clear, colorless colonies.
Procedure:1. Place a membrane filter absorbent pad inside a sterile 60 mm Petri dish
2. Add 1.8-2.0ml Endo Broth to each pad
3. Filter the water sample through a membrane filter
4. Place theb filter top side up on the pad using a rolling motion to avoid entrapping air bubbles
5. Invert the dish and incubate for 22-24 hours at 35±0.5°C
6. Observe and count all colonies that are red and have ametallic sheen.
Expected Results:All colonies that are red and have the characteristic metallic sheen are considered coliforms. The sheen may cover the entire colony, may only be in the center or may appear only around the edges.
Limitations of the Procedure:Occasionally, non-coliform organisms may produce typical sheen colonies. Coliform organisms may also occasionally produce atypical colonies (dark red or nucleated colonies without sheen). It is advisable to verify both colony types (see ref 2).
Certificate of Origin:The raw animal product used in the manufacturing of Endo Broth is derived from casein originating in the USA, New Zealand and/or Australia. The animals used received both ante- and post-mortem inspections under certified veterinary care and were inspected to be free of apparent infectious and contagious diseases. During processing of the raw material, a batch is heated to a minimum of 80°C for 1 hour including a minimum of 5 minutes at 105°C. The product is then dried at a minimum of 140°C.
Important Note: This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.
