In 1943 the author described a test for blood in urine in which small squares of filter paper impregnated with o-tolidine were used. The papers, however, deteriorated within a few months. This disadvantage has now been overcome by using solid o-tolidine hydrochloride instead of the papers.Reagents o-Tolidine hydrochloride. Acid-peroxide solution. Equal volumes of glacial acetic acid and 3 per cent hydrogen peroxide (freshly diluted from 30 per cent). The mixture is allowed to stand for twenty-four hours before using. The solution should be freshly prepared every three months. Procedure A small knife-point (about 3 mg.) of solid o-tolidine hydrochloride is placed on a clean white porcelain tile. One drop of urine is added and thoroughly mixed with the solid by stirring with a glass rod. One drop of acid-peroxide solution is added.ReactionsNegative reaction-A light brown colour slowly develops. Occasionally one or two large blue dots appear. These must be disregarded.Positive reactions.-(a) Numerous small greenish-blue dots develop within a few seconds. Sometimes the dots are drawn out into greenishblue streaks. The dots or streaks persist for periods varying from ten to thirty seconds to several minutes according to the amount of blood present. They then fade and disappear and a light brown colour develops. Reaction (a) is the typical reaction for occult blood (red cells) in urine.(b) When a large amount of blood is present, such as 1,000 red cells per c.mm., a dense, dark blue colour appears immediately. The dark blue mass is surrounded by yellowish-green areas. The colour may persist for half an hour or longer.(c) If free haemoglobin is present a diffuse greenish-blue to dark blue colour appears which persists for a varying -length of time and then fades.False positive reactions.-Bromides and iodides in the urine give positive reactions. These will obscure the reactions for blood in a urine which contains both blood and bromides or iodides. Bromides give a diffuse greenish to greenish-blue colour which develops slowly and then fades. The reactions given by iodides are similar to the blood reactions (b) and (c)
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