Plasma concentrations of glycated hemoglobin (HbA1c; G7 HPLC analyzer, Tosoh Bioscience Inc., South San Francisco, California), fibrinogen (BCS XP System, Siemens Healthcare, Erlangen, Germany), immunoglobulin G (IgG; BN II System, Siemens Healthcare), haptoglobin (Hp; BN II System, Siemens Healthcare) and C-reactive protein (CRP; BN II System, Siemens Healthcare) were measured from blood samples with standard laboratory tests.
Bn 2 system
Manufactured by Siemens
Sourced in Germany, United States
The BN II System is a laboratory equipment product manufactured by Siemens. It is a fully automated, high-throughput system designed for the analysis of blood and other biological samples. The core function of the BN II System is to perform nephelometric and turbidimetric measurements to determine the concentration of specific proteins and other analytes in the samples.
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Lab products found in correlation
N latex rf kit, by Siemens (5 mentions)
Cobas e411, by Roche (5 mentions)
Cobas c711 module, by Roche (4 mentions)
Cardiophase hscrp, by Siemens (4 mentions)
Cobas 6000, by Roche (4 mentions)
Cobas integra 800 cts analyzer, by Roche (3 mentions)
Modular p chemistry analyzer, by Roche (3 mentions)
N latex cdt test, by Siemens (3 mentions)
Xn 1000 hematology system, by Sysmex (2 mentions)
Centaur xp, by Siemens (2 mentions)
Human cystatin c elisa, by BioVendor (2 mentions)
Specific elisa, by Thermo Fisher Scientific (2 mentions)
Coulter lh 780 hematology analyzer, by Beckman Coulter (2 mentions)
Elecsys system, by Roche (2 mentions)
Hlc 723g8, by Tosoh (1 mentions)
Qiaamp dna blood mini kit, by Qiagen (1 mentions)
Streptozotocin (stz), by Merck Group (1 mentions)
Modular e170 analyzer, by Roche (1 mentions)
Microvue creatinine eia kit, by Hitachi (1 mentions)
8 isoprostane, by Abcam (1 mentions)
78 protocols using bn 2 system
1
Biomarker Profiling in Clinical Samples
2
Carbohydrate-Deficient Transferrin Analysis
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CDT, as the percentage of total transferrin (%CDT), was assayed by particle-enhanced imunononephelometry using N Latex CDT test (Siemens Healthcare Diagnostics, Marburg, Germany) on BN II System (Siemens Healthcare Diagnostics, USA). The reference interval of %CDT values ranged from 1.19 to 2.47% (1 st to 99 th percentile). The concentration of transferrin (reference interval: 2.0-3.6 g/L) was determined by the immunonephelometry with specific antibodies (N Antisera to human transferrin and haptoglobin) (Siemens Healthcare Diagnostics Products GmbH, Marburg, Germany) on BN II System. The results were expressed in absolute (mg/L) and relative units (% of total transferrin concentration).
3
Quantifying Plasma RF and IgG
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Plasma RF concentrations were quantitated using the BNII System and N Latex RF
Kit (Siemens Healthcare Diagnostics Inc., Newark, USA). IgG concentrations were
determined using the BNII System and N Latex IgG Kit (Siemens Healthcare
Diagnostics Inc.) according to the manufacturer’s instructions.
Kit (Siemens Healthcare Diagnostics Inc., Newark, USA). IgG concentrations were
determined using the BNII System and N Latex IgG Kit (Siemens Healthcare
Diagnostics Inc.) according to the manufacturer’s instructions.
4
Comprehensive Metabolic Profiling Protocol
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Venous blood samples were collected between 6:00 a.m. and 9:00 a.m. after overnight fasting. FBG, creatinine, total cholesterol, triglyceride, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol (HDL-C) were measured using the Beckman Coulter Chemistry Analyzer AU5400 series (Beckman Coulter Biomedical K.K, Tokyo, Japan). Glycosylated hemoglobin (HbA1c) was measured by high-performance liquid chromatography using Tosoh HLC-723G8 (Tosoh Corporation, Yamaguchi 746-0042, Japan). Screening for UACR was performed using a random, spot urine sample, and the results were expressed in milligrams per gram of creatinine. Urine albumin was measured by nephelometry immunoassay with the SIEMENS BN II System (Siemens Healthcare Diagnostics Products GmbH, 35041 Marburg, Germany).
5
Dried Blood Spot Genotyping for Alpha-1 Antitrypsin Deficiency
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Blood samples were collected with the use of filter paper cards (Whatman 903, lot W101; Whatman/GE Healthcare, Florham Park, NJ, USA). They were transported to the Federal University of São Paulo Hospital São Paulo Central Laboratory, located in the city of São Paulo, Brazil, under temperature-controlled conditions (i.e., at a constant temperature of −20°C), in accordance with applicable International Air Transport Association regulations. All cards were stored at −20°C for subsequent analysis (determination of DBS AAT levels, genotyping, and SERPINA1 gene sequencing). Serum and eluate samples were analyzed on a Siemens BNII system (Siemens Healthcare, Indianapolis, IN, USA) in July of 2012.
For DNA extraction, DBS samples were removed from the cards with a 6-mm paper punch, DNA being extracted with the QIAamp DNA Blood Mini Kit (QIAGEN, Hilden, Germany), in accordance with the manufacturer instructions. For identification of S and Z alleles in exons 3 and 5, respectively, real-time PCR was used with TaqMan(r) SNP Genotyping Assays (Thermo Fisher Scientific Inc., Waltham, MA, USA). All patients with AATD but without S and Z alleles underwent SERPINA1 gene sequencing (exons 2-5) in order to identify other polymorphisms described in the literature.
For DNA extraction, DBS samples were removed from the cards with a 6-mm paper punch, DNA being extracted with the QIAamp DNA Blood Mini Kit (QIAGEN, Hilden, Germany), in accordance with the manufacturer instructions. For identification of S and Z alleles in exons 3 and 5, respectively, real-time PCR was used with TaqMan(r) SNP Genotyping Assays (Thermo Fisher Scientific Inc., Waltham, MA, USA). All patients with AATD but without S and Z alleles underwent SERPINA1 gene sequencing (exons 2-5) in order to identify other polymorphisms described in the literature.
6
Comprehensive Hematology and Biochemistry Analysis
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The Sysmex XN-1000 Hematology System (Sysmex Corporation, Kobe, Japan) was used to perform cell blood count of patients. Biochemical tests were analyzed from serum by spectrophotometric method using Beckman Coulter Olympus AU2700 Plus Chemistry Analyzer (Beckman Coulter, Tokyo, Japan). Ferritin was evaluated by a chemiluminescence immunoassay (Centaur XP, Siemens Healthcare, Germany). Prothrombin time (PT), activated partial prothrombin time (aPTT), and fibrinogen were determined with a fully digital coagulation device of Ceveron-Alpha (Diapharma Group Inc., West Chester, Canada). C-reactive protein (CRP) was measured by the nephelometric method on the BN ™ II System (Siemens, Munich, Germany). Procalcitonin (PCT), D-dimer and Troponin were analyzed from whole blood on the AQT90 flex Radiometer® (Bronshoj, Denmark).
7
Diabetic Nephropathy in Pemt Mice
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Male Pemt−/− and Pemt+/+ mice were housed in cages and maintained on a 12-hour light-dark cycle and received normal chow (MF, Oriental Yeast, Co., Ltd). Eight-week-old mice received five consecutive intravenous injections of 50 mg/kg streptozotocin (STZ) (Sigma, St. Louis, MO) in citrate buffer at pH 4.6 or citrate buffer only (as a control). A blood glucose level over 300 mg/dl was confirmed three days after the STZ administration at two different time points. We sacrificed the mice at 25 weeks of age and subjected them to the subsequent studies. All animal experiments were approved by the Animal Care and Use Committee of the Department of Animal Resources, Advanced Science Research Center, Okayama University.
The serum total homocysteine levels were measured by an enzymatic colorimetric assay (Alfresa Co., Tokyo), the quantification of PC and PE was performed by thin-layer chromatography (TORAY Research Center, Tokyo) and the urinary albumin levels were measured using goat-antiserum against mouse albumin (Cappel) and a turbidimetric immunoassay in the BN II System (Siemens, Eschborn, Germany) and were normalized against the creatinine levels.
The serum total homocysteine levels were measured by an enzymatic colorimetric assay (Alfresa Co., Tokyo), the quantification of PC and PE was performed by thin-layer chromatography (TORAY Research Center, Tokyo) and the urinary albumin levels were measured using goat-antiserum against mouse albumin (Cappel) and a turbidimetric immunoassay in the BN II System (Siemens, Eschborn, Germany) and were normalized against the creatinine levels.
8
Blood Biomarker Analysis Protocol
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Blood samples were collected in the morning after an overnight fast, directly into tubes containing heparin, and centrifuged. Measurements were performed in the clinical chemistry laboratory of the University Medical Center Groningen. Serum levels of total and high-density lipoprotein cholesterol were measured using an enzymatic colorimetric method, triglycerides using a colorimetric UV method, and low-density lipoprotein cholesterol using an enzymatic method, all on a Roche Modular P chemistry analyzer (Roche, Basel, Switzerland). HbA1c was measured using a turbidimetric inhibition immunoassay on a Cobas Integra 800 CTS analyzer (Roche Diagnostics Nederland BV, Almere, Netherlands). Fasting blood glucose was measured using a hexokinase method, and hs-CRP was analyzed using nephelometry (BN II system; Siemens, Marburg, Germany). Results of the thyroid hormone status were available in a subset of 4479 participants. TSH, fT4, and fT3 were assayed by electrochemiluminescent immunoassay on the Roche Modular E170 Analyzer using kits provided by the manufacturer (Roche, Basel, Switzerland) (27) . Normal values are 11-20 pmol/L for fT4 and 4.4-6.7 pmol/L for fT3. The general Dutch population is iodine sufficient. Tests to measure antithyroid peroxidase antibody levels were not performed.
9
Serum IgG and IgG4 Quantification
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Serum levels of immunoglobulin G (IgG) and IgG4 were measured using an immune nephelometric analyzer with the BN™ II System (Siemens, Germany). Human immunoglobulin G4 N latex reagent, L2SUBM, L2PWSM, quality control, and standard products were purchased from Siemens, Germany. Detection was carried out according to the manufacturer's instructions.
10
Complement Levels Measurement Protocol
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The primary exposure variable was complement levels. ELISA assays measuring complement C3 and C4 were performed on EDTA plasma samples according to the manufacturer’s protocol. Low c3 was considered below 90 mg/dL, and low c4 was considered below 10 mg/dL. The complement cut offs were measured according to the manufacturer recommendation (BN™ II System, Siemens Healthineers).
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