Blood urea nitrogen (BUN), serum creatinine, urine creatinine and urine protein were assayed using an automatic analyzer (Beckman Coulter, Inc., Brea, CA, USA). Creatinine clearance and urine protein-to-creatinine ratio were calculated.
Automatic analyzer
Manufactured by Beckman Coulter
Sourced in United States
The Automatic Analyzer is a laboratory instrument designed for rapid and automated analysis of various sample types. It performs a range of analytical tests, including clinical chemistry, immunoassays, and other diagnostic procedures, with high throughput and reproducibility. The core function of the Automatic Analyzer is to automate sample processing, reagent handling, and data analysis to streamline laboratory workflows and enhance efficiency.
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16 protocols using automatic analyzer
1
Kidney Function Biomarker Assessment
2
Plasma Pharmacokinetics and Liver Toxicity
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After the last sample in the plasma pharmacokinetic study, all rats were deeply anesthetized and exsanguinated by severing the abdominal aorta. Blood and liver samples were harvested. The plasma was used for the assay of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), unconjugated bilirubin (IBIL), and conjugated bilirubin (DBIL) by an automatic analyzer (Beckman Coulter, Brea, CA, United States). Liver specimens were fixed in 4% paraformaldehyde for 2–3 days and passed through the routine paraffin embedding procedures. They were serially sectioned and stained with hematoxylin and eosin for histological evaluations.
3
Measuring Serum Biomarkers in Patients
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After an overnight fast of at least 12 hours, a venous blood sample was obtained from the forearm of each patient, as described in detail in our previous study.[24 (link)] Serum Hcy was measured using an enzymatic cycling method. Serum TC, LDL-C, HDL-C, and TGs were measured by enzymatic methods with commercially available kits on a Beckman Coulter Automatic Analyzer. Serum ApoAI and ApoB levels were detected by immunoturbidimetric immunoassays. All of these biochemical analyses were performed at the Clinical Laboratory of Guangxi Academy of Medical Sciences and the People’s Hospital of Guangxi Zhuang Autonomous Region. According to the results from previous studies,[24 (link)] Hcy ≥ 15 μmol/L is often defined as HHcy.
4
Serum and Hepatic Lipid Quantification
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Serum triglycerides and total cholesterol levels were determined using an automatic analyzer (Beckman Coulter, Inc., Brea, CA, USA). Triglycerides and total cholesterol in the liver were measured using colorimetric kits obtained from Abcam, Cambridge, UK (No. ab65336 and ab65390, respectively) according to the manufacturer’s instructions.
5
Renal Function Biochemical Evaluation
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Blood urea nitrogen (BUN), serum creatinine, creatinine clearance, and urine protein-to-creatinine ratio were used as an index of renal function. All biochemical determinations were performed using an automatic analyzer (Beckman Coulter, Inc., Brea, CA, USA). Creatinine clearance was calculated using the standard clearance formula.
6
Plasma CRP and IL-6 Quantification
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C-reactive protein (CRP) and IL-6 levels were determined from plasma per the approved clinical protocols on hospital patients (n = 8 to 12 subjects/group; total subjects = 68). Briefly, CRP in plasma samples was determined by particle-enhanced immunoturbidimetric assay where human CRP agglutinates with latex particles coated with monoclonal anti-CRP antibodies, using the Roche Cobas c-702 analyzer (Roche, Switzerland). The aggregates were determined turbidimetrically, and the analyzer automatically calculated CRP concentration in the sample (milligrams per deciliter) without manual intervention. Likewise, for IL-6 quantification, we used an automatic analyzer (Beckman Coulter, USA) that performs a one-step immunoenzymatic (“sandwich”) assay. Briefly, the plasma sample is placed in a reaction vessel and incubated with paramagnetic particles coated with mouse monoclonal anti-human IL-6, blocking reagent, and the alkaline phosphatase conjugate. After incubation, materials are washed, followed by addition of chemiluminescent substrate. The light generated by the reaction is measured with an inbuilt luminometer. The concentration (picograms per milliliter) of analyte in the sample is determined from a multipoint calibration curve.
7
Biochemical Analysis of Rodent Sera
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The biochemical parameters measured in rodent sera included serum cholinesterase (CHE), ALT, AST, TC, TG, GLU, LDL, and high-density lipoprotein (HDL), using an automatic analyzer (Beckman Coulter, Brea, CA, USA) in accordance with the instructions of the manufacturer. A human-glucocorticoid-enzyme-linked immunosorbent assay (ELISA) Kit (SAB Biotech, College Park, MD, USA) was used to establish a standard curve, to detect intracellular glucocorticoid levels. The absorbance at the corresponding maximum absorption wavelength was measured with a spectrophotometer.
8
Plasma CRP and IL-6 Quantification
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C-reactive protein (CRP) and IL-6 levels were determined from plasma per the approved clinical protocols on hospital patients (n = 8 to 12 subjects/group; total subjects = 68). Briefly, CRP in plasma samples was determined by particle-enhanced immunoturbidimetric assay where human CRP agglutinates with latex particles coated with monoclonal anti-CRP antibodies, using the Roche Cobas c-702 analyzer (Roche, Switzerland). The aggregates were determined turbidimetrically, and the analyzer automatically calculated CRP concentration in the sample (milligrams per deciliter) without manual intervention. Likewise, for IL-6 quantification, we used an automatic analyzer (Beckman Coulter, USA) that performs a one-step immunoenzymatic (“sandwich”) assay. Briefly, the plasma sample is placed in a reaction vessel and incubated with paramagnetic particles coated with mouse monoclonal anti-human IL-6, blocking reagent, and the alkaline phosphatase conjugate. After incubation, materials are washed, followed by addition of chemiluminescent substrate. The light generated by the reaction is measured with an inbuilt luminometer. The concentration (picograms per milliliter) of analyte in the sample is determined from a multipoint calibration curve.
9
Biochemical Parameters Assessment Protocol
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Biochemical parameters were divided into four categories: liver function test (LFT), renal function test (RFT), lipid profile, and electrolytes. LFT included bilirubin, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) and RFT included blood urea and creatinine. LFT and RFT were measured photometrically by using diagnostic kits for DiaSys diagnostic system. Lipid profile included total cholesterol, triglycerides, high-density lipoproteins (HDL), and low-density lipoproteins (LDL). All these parameters were measured using kits provided by Beckman diagnostics on automatic analyzer (Beckman Coulter). Electrolytes included blood sodium and potassium level which were measured by flame photometric method.
10
One-Week Toxicity Evaluation of Di-GLP-1 and Lip-Di-GLP-1
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The one week toxicity test of Di-GLP-1 and Lip-Di-GLP-1 was conducted on db/db mice (n = 3), using a previously described method with some modifications. Mice were divided into five groups, before the testing day each group of mice were fasted overnight, and saline, 500 nmol kg−1 of liraglutide (chemical), liraglutide (biotechnological), and Di-GLP-1 and Lip-Di-GLP-1 were s.c. injected and rodent chow were provided immediately. During the testing period, food and water were provided ad libitum. Tested compound and saline were twice daily administered at 8 AM and 8 PM. At the end of the test, the blood was collected through extracting eyeball (approximately 1 mL) and mice were sacrificed. Blood samples were centrifuged to separate serum. The levels of SCr, AST, BUN, and ALT in serum were determined using Beckman Coulter Automatic Analyzer using commercially available kits.20 (link)
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