New method for the determination of the net bilirubin absorbance in cerebrospinal fluid that minimizes the interference of oxyhaemoglobin and biliverdin

Abstract

The presence of oxyhaemoglobin and biliverdin interferes with the method recommended by the UK NEQAS Specialist Advisory group for EQA of CSF Proteins and Biochemistry for estimating of the net bilirubin absorbance in CSF. This is easily demonstrated by using solutions with different concentrations of these three substances.The two secondary peaks of the oxyhaemoglobin spectrum at 540 nm and 577 nm are used as reference to minimize these interferences. Those peaks have the same absorbance as at 456 nm in the oxyhaemoglobin spectrum, independent of its concentration. This wavelength is very close to the maximum absorption of bilirubin and, therefore, is suitable for estimating the net bilirubin absorbance.A preliminary study with 48 spectrophotometric analyses of CSF from patients who were suspected of having subarachnoid haemorrhage were used to compare both net bilirubin absorbance estimation methods.The new method is practically free of oxyhaemoglobin and biliverdin interference. This allows for higher sensitivity and a more realistic estimation of the bilirubin concentration in a sample.A better estimation of the bilirubin concentration can have special relevance for diminishing the amount of equivocal or inconclusive cases and also to improve the prematurity of the diagnosis.