An Alternative Approach for Detecting Interferences in Enzymatic Acetaminophen Assays

To the Editor: We read with interest the recent description of yet another case of bilirubin-related positive interference in an enzymatic colorimetric assay for acetaminophen (1). As a remedy, these authors suggest measuring bilirubin before every acetaminophen assay and removing it by ultrafiltration when the bilirubin concentration is higher than the reported cutoffs for interference. An alternative and perhaps less costly approach would be to estimate an icteric index, an approximation of the bilirubin concentration based on an absorbance measurement, and to dilute the sample if the icteric index is high. This procedure stemmed from an incident within our laboratory program in which a physician questioned the validity of an acetaminophen result (Roche enzymatic assay) for a patient who had denied taking acetaminophen. In this case, we obtained 4 different samples from this patient (over 4 consecutive days) and measured both bilirubin and acetaminophen in the 4 undiluted samples [bilirubin range, 19.5–23.3 mg/dL (333–399 μmol/L); acetaminophen range, 14–18 μg/mL (93–120 μmol/L)] as well as in serial dilutions (1 volume in 2 and 1 volume in 4 with the therapeutic drug monitoring diluent). The mean (SD) recovery percentages for the bilirubin concentration after the 1-in-2 and 1-in-4 dilutions were 99% (1) and 98% (2), respectively, whereas for acetaminophen the recoveries were only 64% (3) and 38% (4) (Fig. 1). Thus, our experience with the Roche acetaminophen enzymatic assay has been that falsely increased acetaminophen results drop more than expected upon serial dilution, whereas bilirubin results follow the expected linear pattern. Importantly, we have also confirmed the linearity of this acetaminophen assay for samples with high acetaminophen concentrations that do not have this interference (2). These data add to the literature, which suggests that the enzymatic acetaminophen assays are susceptible to a chemical, rather than a spectrophotometric, interference in samples with high bilirubin concentrations (3). Also of note is our simple dilution protocol to detect the presence of a positive interference. For more accurate quantification of acetaminophen, there are interference-free methods [e.g., chromatography, mass spectrometry, enzyme-multiplied immunoassay technique (EMIT)] (4). Given that the presence of positive interference in such enzymatic assays can be quickly uncovered with the icteric index and a dilution protocol, any confirmatory testing can be performed with less urgency. This consideration is important, because the presence of measurable acetaminophen at any concentration can indicate harm, depending on the timing and many other factors (5). In view of the potential for harm, the possible presence of an interference warrants further investigation.

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