Staining Method of Fontana, Spirochetes, silver salt, Ethyl Alcohol, Ammonical Silver Nitrate, Organisms, Fontana Mordant, Reagent

Staining Method of Fontana

	Staining Method of Fontana Spirochetes are very delicate and heat sensitive therefore, they are fixed using chemical fixative in this method. Use of mordant facilitates the precipitation of silver salt. On application of ammonical silver nitrate, it yields a preparation in which the apparent width of the organism is adequately increased and the organisms are seen opaque black against light back ground.
The sequence of steps given by Fontana (3) is as under. Prepare a smear on clean glass slide and air dry. Treat the smear three times with the Fontana's fixative (S52), each for 30 seconds. Rinse the smear with absolute ethyl alcohol and allow it to react for 3 minutes. Drain off the excess of alcohol and dry the slide. Flood the smear with Fontana's mordant (S53) and heat the slide from below till steam rises for 30 seconds.Cool Wash with distilled water and dry the slide. Treat the smear with Fontana's stain, (S54) and heat the slide until steam rises and the smear becomes brown in colour
Wash, dry and examine under oil-immersion objective. Spirochetes will appear as brownish black against yellow background. It may please be noted that unmounted smear tends to fade rapidly and therefore, cannot be stored for more than one day. Moreover, preparation of reagent in this technique is complicated and results vary in quality from day to day