The Wako vanadate oxidation method for bilirubin measurement is unaffected by haemoglobin (≤500mg/dL) or ascorbic acid (≤50mg/dL), both of which can push diazo methods off-target.
New born babies lack the intestinal bacteria that help process bilirubin and neonatal bilirubineamia is a common condition. Typically this resolves itself in a couple of days. However, in some instances, the bilirubineamia is a result of red blood cell destruction caused by new born and maternal blood type incompatibilities or other genetic factors. In these cases it is important that the bilirubin levels do not get too high because excess bilirubin damages developing brain cells in young babies and can cause mental retardation, physical abnormalities or blindness.
Bilirubin in Adults
Bilirubin tests are used to assess liver function in adults. Typically bilirubin is measured alongside other liver disease markers to aid in the diagnosis and monitoring of cirrhosis, hepatitis or gallstones. For example, bilirubin is measured together with hyaluronic acid,α-2-macroglobulin and γ-glutamyl transferase as part of the Hepascore algorithm for liver fibrosis.
The vanadate oxidation method for bilirubin was developed by Wako. Bilirubin is oxidised to biliverdin and the change in absorbance as measured in the yellow part of the spectrum is used to calculate the bilirubin content of the sample. The method is unaffected by haemoglobin (≤500mg/dL) or ascorbic acid (≤50mg/dL), both of which can push diazo methods off-target.
Both the Direct and Total Bilirubin assays utilise the vanadate oxidation principle offering excellent performance with minimal interference to deliver clear, reliable answers when it matters.
Bilirubin is a product of haem catabolism excreted in bile and urine. Small amounts of bilirubin are present in the blood as a result of damaged and dead red blood cells which are disposed of via the spleen. As the erythrocytes are broken down the haeme is turned into unconjugated or indirect bilirubin. Unconjugated bilirubin is water insoluble and is bound to albumin in the blood and transported to the liver. Here the indirect bilirubin is conjugated to glucuronic acid. The resulting conjugated or direct bilirubin is water soluable and much of this goes into the bile and therefore into the small intestine.
The Diazo reaction was described by Ehrlich 1883 and first applied to the determination of bilirubin by van den Bergh and Muller in 1916. Diazo based methods using different chemical accelerators have been widely used ever since. However, Diazo based protocols are prone to negative interference from haemoglobin which presents a danger that results below the true value may be reported. In contrast, high levels of Ascorbic acid can falsely elevate the test result.