Estimating intra- and inter-assay variability in salivary cortisol

Area of Science:

• Endocrinology
• Biochemistry
• Assay Development

Background:

• Intra- and inter-assay coefficients of variation (CVs) are crucial for assessing salivary cortisol enzyme immunoassay (EIA) precision.
• Limited guidance exists on optimal sample selection for CV assessment in salivary cortisol EIA.

Purpose of the Study:

• To compare intra- and inter-assay CVs for salivary cortisol EIA using controls, standards, and unknown samples.
• To evaluate the impact of sample type (fresh vs. frozen saliva) on CVs in salivary cortisol EIA.

Main Methods:

• 174 duplicate samples (controls, standards, unknowns) were analyzed using a commercial salivary cortisol EIA kit.
• Student saliva samples were tested in both fresh and previously frozen states.
• Intra- and inter-assay CVs were calculated and compared using a priori criteria (≤4% intra-assay, ≤7% inter-assay) and a significance level of p < .05.

Main Results:

• Mean intra-assay CVs were ≤2.5% and mean inter-assay CVs were ≤2.8% across all sample types.
• No significant differences in CVs were observed among controls, standards, or unknown samples.
• A statistically significant difference in inter-assay CV was found between fresh and frozen saliva (p = .043), but the difference was not clinically meaningful due to minimal overall error.

Conclusions:

• Salivary cortisol EIA precision can be effectively estimated using controls, standards, or unknown samples.
• Both fresh and frozen saliva samples yield reliable precision estimates for salivary cortisol EIA.
• The study confirms the overall low measurement error of the salivary cortisol EIA kit used.