ENDRASHIKA-KLEGHORNA-GROFA METHOD (L. Jendrassik, R.A. Cleghorn, P. Grof) — a quantitative photometric method of definition of a direct and indirect bilirubin in blood serum.
The principle of a method is that at interaction sulphanilic to - you with sodium nitrite are formed diazofenilsulfonovy to - that (see the Diazo test) edges with bilirubin glucuronide (see. Glucuronides ), so-called direct bilirubin, pink-violet coloring gives blood sera. Determine quantity of direct by intensity of coloring bilirubin (see).
At addition to blood serum of a caffeine reactant untied (indirect) bilirubin turns in soluble (straight line) and decides quantitatively on a diazoreactant. Determine concentration in blood serum of the general bilirubin by intensity of coloring, and between the general and connected (direct) bilirubin determine amount of untied bilirubin by a difference.
Reactants. 1. Caffeine reactant: 5 g of pure caffeine, 7,5 g of sodium benzoate, 12,5 g of sodium acetate (CH 3 COONa • ZN 2 O) dissolve in 90 ml a dist, waters, heat to 50 — 60 °, mix, cool and bring a dist, water to 100 ml. 2. Isotonic solution of sodium chloride. 3. The diazomix prepared just before work and consisting of 10 ml of a diazoreactant I and 0,3 ml of a diazoreactant of II. Diazoreactant of I: 1,5 g sulphanilic to - you dissolve during the heating in 300 — 400 ml a dist, waters, add 15 ml konts. salt to - you with ud. weighing 1,19; after full dissolution sulphanilic to - you and coolings of solution add it a dist, water to 1 l. Store in a dark bottle. Diazoreactant of II: 0,5% solution of sodium nitrite (NaNO 2 ). Store no more than 2 — 3 weeks in a dark bottle.
Course of definition: in 3 test tubes (the general and direct bilirubin — the 1st and 2nd test tubes and control — the 3rd test tube) pour 0,5 ml of the blood serum divorced twice; add 1,75 ml of a caffeine reactant to the 1st and 3rd test tubes, add respectively 1,75 and 0,25 ml of isotonic solution of sodium chloride to the 2nd and 3rd test tubes and then add diazomix on 0,25 ml to the 1st and 2nd test tubes.
For definition of direct bilirubin the kolorimetrirovaniye is carried out at 536 nanometers in 5 — 10 min. after addition of diazomix; for definition of the general bilirubin of test kolorimetrirut in 20 min. against water. Use ditches 0,5 cm wide. Calculation is made according to a standard curve.
Normal blood serum of the person contains 0,5 — 1,2 mg of % of the general bilirubin, 75 its % fall to the share of an indirect bilirubin.
E. — To. — Of m is accepted as the unified a wedge, laboratory method.
Bibliography: Biochemical methods of a research in clinic, under the editorship of A. A. Pokrovsky, page 383, M., 1969; Dzhordzhesku P. and Peunesku E. Biochemical methods of the diagnosis and a research, the lane from Romanians., page 163, Bucharest, 1963; Jendrassik L. u. With leghorn R. A. Photometrische Bilirubinbestim-mung, Biochem. Z., Bd 289, S. 1, 1936, Bibliogr.; Jendrassik L. u. Crof P. Vereinfachte photometrische Methoden zur Bestimmung des Blutbilirubins, ibid., Bd 297, S. 81, 1938.
B. A. Popkov.