Ilab 300 plus

Hematological analysis was performed in the blood samples collected into K-EDTA tubes (Greiner Bio-one, Kremsmünster, Austria), using an automated hematology analyzer Cell-Dyn 3700 (Abbott Diagnostics, Illinois, USA). To examine biochemical parameters, blood samples were drawn from an antecubital vein into sample tube with serum separator gel (Greiner Bio-one, Kremsmünster, Austria) and then left to coagulate for 30 min to separate the serum, and the obtained samples were stored at −80°C for later analysis. The following items were included in the general biochemical examination: aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatine kinase (CK), lactate dehydrogenase (LDH), uric acid (UA), creatinine (Cre), high sensitivity C-reactive protein (hs-CRP), total cholesterol (CHOL), HDL cholesterol (HDL-C), and triglycerides (TG). Biochemical analysis was performed using an ILab 300 Plus autoanalyser (Instrumentation Laboratory, Milan, Italy) employing commercial kits (Bioanalytica, Belgrade, Serbia). The concentration of LDL cholesterol (LDL-C) was calculated using the formula of Friedewald et al. [22 (link)].

Briefly, blood samples were centrifuged for 15 min at 3,500 rpm (SIGMA 2–5 Centrifuge, Osterode am Harz, Germany) and the serum was separated. Blood urea nitrogen (BUN), creatinine, and alanine aminotransferase (ALT) concentrations were measured using an ILab-300 Plus clinical chemical analyzer and commercial kits (both from Instrumentation Laboratory, Bedford, MA, USA). The levels of these substances were measured to exclude possible toxic effects of the J. mollis extract on the liver and kidney and to assess its effects on renal and hepatic function.

The concentrations of creatinine and urea in serum were determined using a modified Jaffe colorimetric method and the urease enzymatic method, respectively, by means of commercial kits and spectrophotometry with an automatic analyser ILAB-Aries (ILab-300 Plus; Instrumentation Laboratory, Bedford, MA, USA).
The serum levels of pro-inflammatory cytokines IL-1β, IL-6 and TNF-α were determined using a sandwich enzyme-linked immunoassay protocol development kit (Peprotech, Mexico City, Mexico). The optical density was measured at 405/620 nm using a microplate-reading spectrophotometer (Mutilskan FC, Thermo Fischer Scientific Oy, Vantaa, Finland). The results are expressed as ng/mL.
For quantitative detection of rat serum NGAL and KIM-1, Abcam one-step ELISA kits were used (Lipocalin-2/NGAL Rat ELISA Kit, Abcam 119602, Abcam, Cambridge, UK; KIM-1/TIM-1 Rat ELISA Kit Abcam 119597, Cambridge, MA, USA, respectively). Cys-C was measured using ELISA kits from R&D Systems (Quantikine ELISA; Mouse/Rat Cystatin C Immunoassay, R&D Systems, Minneapolis, MN, USA).