I stat portable clinical analyzer

Whole blood was collected from the mice, and the whole blood samples were analyzed immediately on the spot using a portable clinical analyzer, i-STAT Portable Clinical Analyzer (Abbott Laboratories, Abbott Park, IL, USA). A minimum of 100 µl of blood was collected per mouse, and 100 µl of whole blood was loaded into the i-STAT. i-STAT was used to determine the creatinine level in the whole blood.

Blood samples were analyzed using the Abbott i-STAT portable clinical analyzer (Abbott Point-of-Care) and iSTAT 8+ cartridges (Abbott Laboratories) to determine sodium (Na), potassium (K), chloride (Cl), TCO₂, anion gap, ionized calcium (iCa), BUN, creatinine, glucose (Glu), hematocrit (Hct), and hemoglobin (Hb).
Urine albumin was determined using a rat albumin–specific ELISA (Bethyl Laboratories), and creatinine was determined by reagent set (Diazyme, DZ072b-KY1), per the manufacturers’ protocols. The plasma renin sample was measured using ELISA (MilliporeSigma, RAB1162-1KT) according to the manufacturer’s protocol.

At designated time points after burn injury, the animals were anaesthetized with Ketamine and Diazepam (75 and 10 mg/kg, resp., i.p.), the carotid artery was cannulated, and 0.3 mL of heparinized arterial blood was collected for blood gases analysis using the i-STAT Portable Clinical Analyzer (Abbott Laboratories Inc., NJ, USA). Blood parameters, lactate, blood urea nitrogen (BUN), and pH were measured. Three milliliters of blood with EDTA were also collected via the carotid artery for other analyses, with fluid replacement using heparinized saline, shortly before perfusion with saline. The collected blood was centrifuged and clean plasma was aliquoted and stored in −80°C for future assays. The animals were then perfused via transcardial perfusion with saline until the lungs, liver, and kidneys were clear of blood. The required organs were removed and soaked in 10% buffered formalin for fixation, prior to histological examination.

The i-STAT Portable Clinical Analyzer and i-STAT G7+ cartridges (Abbott Point of Care, Chicago, IL) were used. Arterial blood was sampled from mice's left ventricle (n = 6 in each group). It was then introduced into the sample well and allowed to fill by passive movement to the indicated level (80-100 μL). After closing the cap on the model well, the cartridge was inserted into the analyzer. After completing the calibration and analysis cycles successfully, partial arterial oxygen pressure (PaO₂) values were recorded.

Oxygen and air (15%:85%) were supplied through a ventilator (SAR‐1000, CWE, Ardmore, OK for experiments in all mechanical ventilation conditions and spontaneous breathing condition experiments in optical intrinsic signal [OIS] imaging, or TOPO, Kent Scientific, Torrington, CT for the fMRI studies with spontaneous breathing). For spontaneous breathing, gases were supplied through a tube or nose cone with a mask (OIS imaging and fMRI, respectively) at 1 L/minute. In the mechanical ventilation case, exactly the same tubing length (4.45 m, ID: 1.6 mm) was used for both the OIS and fMRI studies. To identify the proper ventilator settings for maintaining normal physiological conditions, extensive preliminary studies were performed with blood gas analysis (i‐STAT Portable Clinical Analyzer; Abbott Point of Care, Princeton, NJ). The determined ventilator settings were an inspiration/expiration ratio of 35%:65%, a respiratory rate of 160 (ketamine/xylazine) or 90 (isoflurane) breaths per minute to match with spontaneous breathing rate. A tidal volume was set to obtain normal blood gas values; 0.0103 × body weight (g) + 0.4117 (i.e., 0.62‐0.7 ml) for ketamine/xylazine, or 0.0366 × body weight (g) + 0.1095 (i.e., 0.91‐1.14 ml) for isoflurane.