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A modification of the acid diazo coupling method (Malloy-Evelyn) for the determination of serum total bilirubin

1997; Taylor & Francis; Volume: 57; Issue: 3 Linguagem: Inglês

10.3109/00365519709060037

1502-7686

Markku Parviainen,

Protein Interaction Studies and Fluorescence Analysis

AbstractA simple and reliable method for the determination of total bilirubin from human serum is described. In this method, indirect bilirubin is liberated by the tenside in 0.12 mol T−1 HC1 (Rl), and the total bilirubin is coupled with a 2,5-dichlorobenzene diazonium (DBD) salt to obtain the corresponding azobilirubin having a λmax of about 520–522 nm. The method can easily be applied to the KONE Delta, a fully automated, discrete random access clinical analyser, and also to less modern instruments. A sample volume of 5 μl, Rl volume of 180 μ1, and R2 volume of 36 μl was used on the KONE Delta. After a 5-min incubation at 37°C, measurement at 575 nm was done (main wavelength). The within-run imprecision (CV%) varied from 2.9 to 0.3% within the serum total bilirubin range of 14–290 (imol 1 (n=10). The between-run imprecision was from 2.2 to 1.3% within the range 13–97 μmol I1 (n=8). The method is linear up to at least 340 μmol T−1 (19.8 mg dl−1), and dilution extends the test limit to 3400 μmol I−1 (198.8 mg dr1). The linearity of dilution was good over the practical measuring range. The present method had a strong linear correlation with the Boehringer 2,5-dichlorophenyl diazonium (DPD) method on the Hitachi 717 analyser: y(DBD) = 1.018x(DPD)+0.758, r=0.9955 (n=61). The stability of R2 (diazo reagent) in the analyser reagent compartment lasts at least 2 weeks.Key Words: azobilirubinbichromatic spectrophotometryblood chemical analysisclinical chemistrydiagnosisintermethod comparison Additional informationNotes on contributorsM. T. ParviainenJoyce Laing works in the Department of Child and Family Psychiatry, Playfield House, Cupar, Fife, and is a Consultant Art Therapist to Psychiatric Hospitals and Prisons and Chairwoman of the Scottish Society of Art and Psychology.