Measurement of late-night salivary cortisol with an automated immunoassay system

Background: Measurement of late-night salivary cortisol
concentrations is increasingly used as a screening test in
suspected Cushing's syndrome. Cortisol concentrations are typically
extremely low in late-night samples and discordant assay-specific
reference ranges have been reported. Therefore, the aim of our
study was to assess the analytical performance of the first
automated cortisol immunoassay specified for salivary measurements
and to establish late-night sampling reference-range data for this
test. Methods: Salivary cortisol was measured using the Roche Cobas
Cortisol assay (Roche Diagnostics). Five salivary pools in
different concentration ranges were used to assess the inter-assay
imprecision of this test in a two-centre evaluation protocol
including two reagent lots. Linearity was tested by serial
dilution. Salivary samples were obtained at 23:00 h from 100
apparently healthy volunteers using a commercially available
salivary sampling device (Salivette, Sarstedt). A subset of 20
samples was used for method comparison with isotope dilution liquid
chromatography-tandem mass spectrometry. Results: Inter-assay
coefficients of variation (n=20) between 11.6% and 40.4% were found
for mean cortisol concentrations between 12.9 and 2.6 nmol/L, with
an estimated functional sensitivity of approximately 5.0 nmol/L.
The test also gave linear results in the lowest concentration range
between 1.0 and 8.3 nmol/L. Mean late-night salivary cortisol of
5.0 nmol/L was found for healthy individuals; the absolute range
was 1.4-16.7 nmol/L, and the 95th percentile was 8.9 nmol/L.
Substantially lower concentrations were found with isotope dilution
LC-MS/MS compared to immunoassay results (mean concentrations 1.8
and 4.4 nmol/L, respectively). Conclusions: The automated assay
investigated was found to offer acceptable analytical performance
in the very low concentration range required for late-night
salivary cortisol, despite a very short turnaround time. Using this
assay, late-night salivary cortisol concentrations below 8.9 nmol/L
are typically found in healthy volunteers.