Serum TSH was analyzed by sandwich chemiluminescence immunoassay (reference range 0.35–4.0 mU/L, CV 19.6% at 0.29 mU/L). Free thyroxine (reference range 11.5–22.7 pmol/L, CV 23% at 11.7 pmol/L) and total triiodothyronine (reference range 1.0-2.6 nmol/L, CV 29.6% at 1.07 nmol/L) were analyzed by competitive immune analysis assay using chemiluminescence technology on ADVIA Centaur XP from Siemens (Siemens Healthcare GmbH). Plasma glucose was analyzed bedside on YSI 2900 Biochemistry analyzer (Xylem Inc. White Plains, NY, USA). Plasma C-peptide was measured using a two-sided electrochemiluminescence immunoassay (Roche Diagnostics). Total ghrelin was measured by radioimmunoassay (RIA) (Millipore). Serum insulin was measured using two-sided electrochemiluminescence (Roche/Hitachi Modular Analytics, Roche Diagnostics). Plasma leptin was analyzed on Human Leptin Immunoassay from R&D Systems (Bio-Techne Corporation, Minneapolis, MN, USA). The concentration of CCK in plasma was measured using an in-house RIA, antibody no. 92128 (16 (link)). Plasma gastrin was measured using an in-house RIA (antibody no. 2604) as previously described (17 (link), 18 (link)). Plasma GLP-1 was measured using antiserum no. 89390 (19 (link)). Plasma acetaminophen was analyzed using enzymatic determination and absorption photometry, Siemens Atellica (Siemens Healthcare).
Atellica
Manufactured by Siemens
Sourced in Germany, United States
The Atellica is a laboratory equipment product offered by Siemens. It is designed to perform various analytical tests and procedures in a clinical or research setting. The core function of the Atellica is to provide accurate and reliable data to support medical decision-making and research activities.
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20 protocols using atellica
1
Comprehensive Biomarker Analysis in Clinical Samples
2
Evaluating Cardiac Troponin Assays Performance
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To evaluate the generalizability of risk stratification thresholds we used stored samples from a substudy of the trial to compare the performance of different high-sensitivity cardiac troponin I assays (Abbott ARCHITECTSTAT and Siemens Atellica, Siemens Healthineers) and high-sensitivity cardiac troponin T (Roche Elycsys, Roche Diagnostics). Participants provided informed consent for additional blood sampling and storage, as described previously.20 (link)–22 (link) The analysis population was defined in the substudy using the same inclusion and exclusion criteria as for the trial population.
3
Comparative Analysis of Cardiac Troponin I
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Two different analytical systems were applied, with analysis being performed in two different centers.
Center 1. Policlinico of Milan. The analyses were performed on the Sgx Clarity System (Singulex, Alameda, CA, USA), using EDTA plasma as sample material. All samples were measured in duplicate, with all samples obtained from the same subject being measured using a single reagent pack in a single analytical run. The same single reagent lot was used for analysis of samples from all subjects. Due to the analytical throughput of the analyzer, a maximum of eight subjects per day were measured (160 measurements/day). Sgx Clarity cTnI Controls (Singulex, Alameda, CA, USA) were used as quality control materials.
Center 2. Hospital Universitario La Paz Madrid. The analyses were performed on Siemens Atellica (Siemens Healthineers, Erlangen, Germany), using serum as sample material. All samples were measured in duplicate. All samples obtained from the same subject were measured using a single reagent pack in a single analytical run. All samples were measured for 3 consecutive days (30 subjects, 600 measurements/day) with the same reagent lot. Liquichek Cardiac Markers Plus (Bio-Rad, Hercules, CA, USA) was used as quality control material.
The technical characteristics of the two analytical methods are provided in Supplementary Table 3.
Center 1. Policlinico of Milan. The analyses were performed on the Sgx Clarity System (Singulex, Alameda, CA, USA), using EDTA plasma as sample material. All samples were measured in duplicate, with all samples obtained from the same subject being measured using a single reagent pack in a single analytical run. The same single reagent lot was used for analysis of samples from all subjects. Due to the analytical throughput of the analyzer, a maximum of eight subjects per day were measured (160 measurements/day). Sgx Clarity cTnI Controls (Singulex, Alameda, CA, USA) were used as quality control materials.
Center 2. Hospital Universitario La Paz Madrid. The analyses were performed on Siemens Atellica (Siemens Healthineers, Erlangen, Germany), using serum as sample material. All samples were measured in duplicate. All samples obtained from the same subject were measured using a single reagent pack in a single analytical run. All samples were measured for 3 consecutive days (30 subjects, 600 measurements/day) with the same reagent lot. Liquichek Cardiac Markers Plus (Bio-Rad, Hercules, CA, USA) was used as quality control material.
The technical characteristics of the two analytical methods are provided in Supplementary Table 3.
4
Serum Biomarkers for Hepatic Steatosis
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The separated sera were stored at 2 °C–8 °C for a maximum of 1 day, then assayed for the 10 serum biomarkers that are included in the FibroMax score. The obtained results were adjusted for age, gender, weight, and height.
Nephelometry from serum samples was used to assess α2-macroglobulin, haptoglobin, apolipoprotein A1, while spectrophotometry from serum samples was used to assess total bilirubin, gamma-glutamyltransferase (GGT), aspartate aminotransferase (AST), alanine aminotransferase (ALT), total cholesterol, and triglycerides. Moreover, plasma fasting glucose was assessed using NaF/K2 oxalate spectrophotometry. The parameters were assayed using BN ProSpec System from Siemens for nephelometry and Siemens Atellica from Siemens for spectrophotometry.
The results of the measured blood variables were entered into the BioPredictive network where the algorithms were computed. In this study, SteatoTest which is considered a measure of the steatosis grade in hepatocytes that ranges from S0–S3, was used in our study in combination with ultrasonography to confirm hepatic steatosis [34 ].
Nephelometry from serum samples was used to assess α2-macroglobulin, haptoglobin, apolipoprotein A1, while spectrophotometry from serum samples was used to assess total bilirubin, gamma-glutamyltransferase (GGT), aspartate aminotransferase (AST), alanine aminotransferase (ALT), total cholesterol, and triglycerides. Moreover, plasma fasting glucose was assessed using NaF/K2 oxalate spectrophotometry. The parameters were assayed using BN ProSpec System from Siemens for nephelometry and Siemens Atellica from Siemens for spectrophotometry.
The results of the measured blood variables were entered into the BioPredictive network where the algorithms were computed. In this study, SteatoTest which is considered a measure of the steatosis grade in hepatocytes that ranges from S0–S3, was used in our study in combination with ultrasonography to confirm hepatic steatosis [34 ].
5
FibroMax Biomarker Evaluation Protocol
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Sera were separated and stored at 2 °C–8 °C for 1 day at most. Afterward, they were assayed for the ten serum biomarkers included in the FibroMax score. Adjustments for age, gender, weight and height of the achieved results were performed for obtaining the final score.
The serum levels of α2-macroglobulin, haptoglobin, apolipoprotein A1 were evaluated using nephelometry (BN ProSpec System from Siemens), while total bilirubin, gamma-glutamyltransferase (GGT), aspartate aminotransferase (AST), alanine aminotransferase (ALT), total cholesterol and triglycerides were assessed using spectrophotometry (Atellica from Siemens). Moreover, NaF/K2 oxalate spectrophotometry was used to assess plasma fasting glucose levels. Obtained values of the evaluated blood variables were completed into the BioPredictive network, where computed algorithms were performed.
The serum levels of α2-macroglobulin, haptoglobin, apolipoprotein A1 were evaluated using nephelometry (BN ProSpec System from Siemens), while total bilirubin, gamma-glutamyltransferase (GGT), aspartate aminotransferase (AST), alanine aminotransferase (ALT), total cholesterol and triglycerides were assessed using spectrophotometry (Atellica from Siemens). Moreover, NaF/K2 oxalate spectrophotometry was used to assess plasma fasting glucose levels. Obtained values of the evaluated blood variables were completed into the BioPredictive network, where computed algorithms were performed.
6
Hormonal assessment during clomiphene citrate therapy
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Study primary outcome was hormonal assessment at baseline and at 1, 3, 6 and 12 months during CC therapy and annually thereafter measured with immunoassay (Atellica®, Siemens, Erlangen, Germany) the immunoassay was replaced in 2014, however, hormonal values were comparable using both immunoassay tools:
Total testosterone (TT) from early morning blood draws (<11 am) (laboratory standard (lab. st.): 7.0–31 nmol/L)
Free testosterone (FT) from early morning blood draws (<11 am) (lab. st.: 230–600 pmol/L)
LH (lab. st.: 1.0–9.0 IU/L)
FSH (lab. st.: 1.0–19.0 IU/L)
Estradiol (lab. st.: 70–200 pmol/L)
Sex‐hormone binding globulin (SHBG) (lab. st.: 10–40 nmol/L)
Albumin (lab. st.: 35.0–50.0 g/L)
Prolactin (lab. st.: 0.10–0.65 IU/L)
7
Comparative Assessment of TSH Assays
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TSH was measured in one run using the automated electrochemiluminescence immunoassays of the following platforms: Cobas® (Roche Diagnostics) Elecsys TSH REF 08429324 190, Alinity® (Abbott Laboratories) TSH reagent kit G71292R02, and Atellica® (Siemens Healthcare Diagnostics) TSH3-UL REF 10995703. The Cobas and Alinity assays are standardized against the same WHO Second International Standard for human TSH (IRP 80/558) and the Atellica assay is standardized against the WHO Third International Standard for human TSH (IRP 81/565). The samples were included based on the initial measurement using the Cobas®. For the method comparison, all the samples were remeasured on the Cobas® platform in one run.
8
Progesterone Measurement in Blastocyst Transfer
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Blood sampling was carried out in a standardized way on theblastocyst transfer day (9 am to 11 am ) 2–4 h after vaginal progesteroneadministration. Pregnancy testing was performed 9–11 days after ET at a random time for patient convenience and serum P4 was included in the analysis. Serum P4 levels were analysed using direct chemiluminescent technology (Atellica, Siemens), routinely used for analysis at the local department of biochemistry. All measurements were performed according tothe manufacturer’s instructions. The assay provided resultsfrom 1.0 to 1908 nmol/l and was designed to have a within-laboratory precision of ±12% (2 Coefficient of Variation) for samples level 6.3 nmol/l and ±8% (2 CV) at samples level 39 nmol/l.
All blood samples were analysed for progesterone immediately.
All blood samples were analysed for progesterone immediately.
9
Comparative Evaluation of Automated fT4 Assays
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Serum fT4 concentrations were measured using five different commercially available automated competitive immunoassays: Alinity (Abbott), Atellica (Siemens), Cobas (Roche; Gen III), Lumipulse (Fuijrebio) and UniCel DXI (Beckman Coulter). Two of these assays were one-step immunoassays (Atellica and Lumipulse) and three of these assays were two-step immunoassays (Alinity, Cobas and UniCel DXI). Albumin concentrations were measured using the Cobas (Roche) colorimetric assay (bromocresol purple method). Serum TBG concentrations were measured using an RIA (Thermo Fischer Scientific). Analyses using Alinity, Atellica, Cobas, Lumipulse and RIA were performed in batch at the Endocrine Laboratory of Amsterdam UMC. Analyses using UniCel DXI were performed in batch at the laboratory of Red Cross Hospital (RKZ) Beverwijk.
10
Serum Biomarkers Evaluation for FibroMax
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We extracted the sera and stored them for a maximum of 1 day at 2°C – 8°C, after which the sera were assayed for the ten serum biomarkers included in the FibroMax score. The obtained result values were inputted into the BioPredictive network, where adjustments for age, gender, weight, and height were performed, along with computed algorithms. Afterward, the final score was obtained for each subject.
Nephelometry (BN ProSpec System from Siemens) was used to assess the serum haptoglobin, apolipoprotein A1, and α2-macroglobulin levels. Spectrophotometry (Atellica from Siemens) was used to assess aspartate aminotransferase (AST), alanine aminotransferase (ALT), total bilirubin, gamma-glutamyltransferase (GGT), triglycerides, and total cholesterol levels. Furthermore, NaF/K2 oxalate spectrophotometry was used to measure plasma fasting glucose levels.
Nephelometry (BN ProSpec System from Siemens) was used to assess the serum haptoglobin, apolipoprotein A1, and α2-macroglobulin levels. Spectrophotometry (Atellica from Siemens) was used to assess aspartate aminotransferase (AST), alanine aminotransferase (ALT), total bilirubin, gamma-glutamyltransferase (GGT), triglycerides, and total cholesterol levels. Furthermore, NaF/K2 oxalate spectrophotometry was used to measure plasma fasting glucose levels.
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