Human saliva total protein levels by AV17 pigment based analysis: validation, stability and short-term variation studies.

BACKGROUND: A colorimetric method based on acid violet pigment, namely AV17, to analyse salivary total protein content, was assessed. METHODS: Human saliva sample or standard (50 microL) was added to 1.5 mL of AV17 working solution (1 mg/mL in 75 mmol/L sodium chloride and 1.7 mol/L phosphoric acid). Total protein concentration was measured at 546 nm. Salivary total protein of healthy subjects was analyzed. RESULTS: The standard protein was Human Serum Albumin and the detection range was 38 mg/L - 900 mg/L with a LOD and LOQ of 26 mg/L and 64 mg/L, respectively. Intraday CVs were 3% - 5% and interday CVs were 3%-6%. The dilution test demonstrated a correlation coefficient of 0.999 and the recovery tests ranged from 108% to 111%. Saliva sample stability was also demonstrated. No intra-individual salivary total protein variation was found during the morning. CONCLUSIONS: The method suitability for laboratory diagnostic purposes to analyse human saliva protein content and stability was demonstrated.

Cortisol assays and diagnostic laboratory procedures in human biological fluids.

The overview of cortisol physiology, action and pathology is achieved in relation to the hypothalamic-pituitary-adrenal axis alteration by laboratory investigation. The measurements of cortisol and related compound levels in blood, urine and saliva used to study the physiological and pathological cortisol involvement, are critically reviewed. The immunoassay and chromatographic methods for cortisol measurement in the various biological fluids are examined in relation to their analytical performances, reference ranges and diagnostic specificity and sensitivity. Moreover, blood, urine and saliva cortisol level measurements are described taking into account the diagnostic implications. The deduction is that each method requires the definition of its own reference range and its related diagnostic cut-off levels. Thus, this review, stressing the analysis procedures, could help to understand and compare the results of the different assays.

Human saliva cortisone and cortisol simultaneous analysis using reverse phase HPLC technique.

BACKGROUND: Hyper-hypo tension (like Cushing's syndrome, apparent mineralocorticoid excess syndrome and Addison's disease) diagnostic laboratory requires cortisol (F) analysis. The simultaneous analysis of human saliva F and cortisone (E), the inactive F metabolite, by solid phase extraction and RP-HPLC was studied. METHODS: Saliva/standard samples were C18-SPE extracted, dried and resuspended. E and F were analysed by isocratic RP-HPLC (acetonitrile/water 27/73%) and UV detection. In the morning and in the evening Salivette stimulated saliva specimens were collected from healthy volunteers. RESULTS: The E and F calibration curve ranges were 11.0-110.0 and 5.5-55.0 nmol/l respectively. The LOD was 0.2 and 0.1 nmol/l for E and F respectively. The intra and inter assay CVs were respectively 2.7-6.6 and 5.6-7.0% for E and 5.8-7.0 and 11.7-13.1% for F. The E and F spiked saliva sample recovery was 99% and 88% respectively. Saliva specimen stability was validated. E and F saliva levels in healthy volunteers were significantly (p<0.001) higher at 8 a.m. compared with 11 p.m. (26.4+/-8.9 vs. 4.3+/-2.9 nmol/l for E; 11.1+/-4.0 vs. 2.5+/-1.5 nmol/l for F, respectively). CONCLUSIONS: This method is suitable for periodic analyses in a clinical biochemistry laboratory for endocrinology investigation purposes, simultaneously analysing E and F levels in a saliva specimen.