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Comparison of Liquid Chromatography Mass Spectrometry and Enzyme-Linked Immunosorbent Assay Methods to Measure Salivary Cotinine Levels in Ill Children


Comparison of Liquid Chromatography Mass Spectrometry and Enzyme-Linked Immunosorbent Assay Methods to Measure Salivary Cotinine Levels in Ill Children

E Melinda Mahabee-Gittens et al. Int J Environ Res Public Health.


Objective: Cotinine is the preferred biomarker to validate levels of tobacco smoke exposure (TSE) in children. Compared to enzyme-linked immunosorbent assay methods (ELISA) for quantifying cotinine in saliva, the use of liquid chromatography tandem mass spectrometry (LC-MS/MS) has higher sensitivity and specificity to measure very low levels of TSE. We sought to compare LC-MS/MS and ELISA measures of cotinine in saliva samples from children overall and the associations of these measures with demographics and TSE patterns. Method: Participants were nonsmoking children (N = 218; age mean (SD) = 6.1 (5.1) years) presenting to a pediatric emergency department. Saliva samples were analyzed for cotinine using both LC-MS/MS and ELISA. Limit of quantitation (LOQ) for LC-MS/MS and ELISA was 0.1 ng/ml and 0.15 ng/ml, respectively. Results: Intraclass correlations (ICC) across methods = 0.884 and was consistent in sex and age subgroups. The geometric mean (GeoM) of LC-MS/MS = 4.1 (range: < LOQ - 382 ng/mL; 3% < LOQ) which was lower (p < 0.0001) than the ELISA GeoM = 5.7 (range: < LOQ - 364 ng/mL; 5% < LOQ). Similar associations of cotinine concentrations with age ( < -0.10, p < 0.0001), demographic characteristics (e.g., income), and number of cigarettes smoked by caregiver ( > 0.07, p < 0.0001) were found regardless of cotinine detection method; however, cotinine associations with sex and race/ethnicity were only found to be significant in models using LC-MS/MS-derived cotinine. Conclusions: Utilizing LC-MS/MS-based cotinine, associations of cotinine with sex and race/ethnicity of child were revealed that were not detectable using ELISA-based cotinine, demonstrating the benefits of utilizing the more sensitive LC-MS/MS assay for cotinine measurement when detecting low levels of TSE in children.

Keywords: ELISA; cotinine; liquid chromatography; secondhand smoke exposure and children.

Conflict of interest statement

The authors declare no conflicts of interest.


Figure 1
Figure 1
Distribution plot of ln-transformed cotinine in saliva (n = 203) detected by LC-MS/MS (blue) and ELISA (red).
Figure 2
Figure 2
Scatter plot of LC-MS/MS vs. ELISA based ln-transformed cotinine measurements > LOQ (N = 203), with a linear regression line for the overall sample. Parameter estimates are provided in inset.

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