Cobas 8000 modular

Intracardial blood samples were centrifuged to separate the serum and frozen at −80 °C for the analysis of biomarkers and the acute phase response. The serum samples were thawed and centrifuged for 10 min at 2,000 g at room temperature. The supernatant was then used for the analyses of aspartate aminotransferase (AST), alanine aminotransferase (ALT), cholesterol, triglycerides and albumin concentrations, performed on a Cobas 8000 modular analyser (Roche, USA).

Clinical and laboratory data during hospital admission were extracted from the electronic medical records, including age, sex, weight, body temperature, respiratory rate, heart rate, systolic blood pressure (SBP), diastolic blood pressure (DBP), the length of hospital stay, PCT, CRP, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and albumin (ALB), creatinine (CREA) and urea nitrogen (UREA). The PCT levels were detected using Cobas^® 8000 modular analyzer (Roche Diagnostics, Switzerland). The CRP levels were detected using a latex-enhanced immunoturbidimetric assay on an UPPER analyzer (Ultrasensitive CRP kit, Upper Bio-Tech, China). The ALT, AST, ALB, CREA, and UREA levels were recorded by the conventional clinical analytical approach using an automatic Beckman biochemical analyzer (Beckman Coulter, Brea, California, USA). The total white blood cells, neutrophils, lymphocytes, and platelets were recorded using an automated blood cell counter (Sysmex Corporation, Japan). In our data analysis, PCT levels>100 ng/mL or <0.02 ng/mL were defined as 101 ng/mL and 0.01 ng/mL, respectively, and CRP levels <0.8 mg/L were defined as 0.7 mg/L. The LCR was calculated as the total lymphocyte count (×10⁹ cells/L)/CRP (mg/L).

On the study day, total cholesterol, triglycerides, and high-density lipoprotein cholesterol were measured on the Roche Cobas 8000 Modular. Approximately 2 weeks before the study day, fasting serum was withdrawn to screen for baseline factors. Dehydroepiandrosterone sulfate (catalog #L2KDS2) was measured using a solid-phase competitive chemiluminescent enzyme immunoassay with an Immulite 2000 XPi system from Siemens Healthcare diagnostics (The Hague, The Netherlands). High-sensitivity C-reactive protein (hsCRP) (catalog #04628918190) was determined by a particle-enhanced turbidimetric assay using Cobas Integra 800 from Roche Diagnostics. Interleukin 6 (catalog #SS600B) and TNF-α (catalog #SSTA00D) were measured by ELISA from R&D Systems. All measurements were performed at the Department of Clinical Chemistry and Laboratory Medicine of the Leiden University Medical Center in The Netherlands.

Collection method: A sterile cotton ball was swabbed inside the mouth for at least 2 min and then placed in a 20-mL sterile syringe. The saliva (2 mL) was transferred to a test tube by pressing the plunger of the syringe. The saliva was centrifuged for 10 min at 3000 rpm, and 1 mL of the sample was stored at −20°C for later analysis. We commissioned Xinjiang Di’An Testing Company to analyze the samples. The chemiluminescence method was used for detection using a Cobas® 8000 modular analyzer (Roche Diagnostics, USA). A cortisol detection kit was also used.