Serum immunoglobulins were measured at diagnosis, during steroid treatment, and at disease relapse within the IgG4-RD cohort. Comparisons among IgG4-RD patients, DC, and HC were made at diagnosis before steroid initiation. Total serum IgG and IgG4 were measured by nephelometry (BNII analyser, Siemens, Oxford, UK). Total IgE was measured by the automated Immunocap method (Phadia 250, Milton Keynes, UK). Elevated serum IgG (≥16 g/L), IgG4 (≥1.4 g/L), and IgE (≥125 kIU/L) were defined by institution range.
Bnii analyser
Manufactured by Siemens
Sourced in United Kingdom
The BNII analyser is a laboratory instrument designed for the analysis of various biological samples. It functions as an automated system for the detection and quantification of specific analytes or substances within a sample. The core purpose of the BNII analyser is to provide accurate and reliable analytical data to support clinical decision-making and research activities.
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Lab products found in correlation
Human leptin quantikine elisa kit, by R&D Systems (1 mentions)
Cyto stat tetrachrome, by Beckman Coulter (1 mentions)
Anti cd62l antibody, by BD (1 mentions)
Advia 1650 analyser, by Siemens (1 mentions)
Fc500 flow cytometer, by Beckman Coulter (1 mentions)
Diamat analyzer, by Bio-Rad (1 mentions)
Tgl flex, by Siemens (1 mentions)
8 protocols using bnii analyser
1
Serum Immunoglobulin Levels in IgG4-RD
2
Geriatric Rehabilitation: Immunological Profiles
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All laboratory variables were assessed upon admission to the geriatric rehabilitation unit. Serum leptin levels were assessed using a commercially available ELISA (Human Leptin Quantikine ELISA Kit (R&D Systems, Abingdon, UK)). The serum samples were stored at −20 °C prior to analysis. The serum albumin was measured using a BNII analyser (Siemens, Saint-Denis, France). C-reactive protein (CRP) was measured by immunoturbidimetry using an Advia 1650 analyser (Siemens Healthcare, Saint-Denis, France). An ultrasensitive CRP assay was not available at the time of the study. Immunological variables were assessed using flow cytometry immunophenotyping, as described previously [25 (link)]. Absolute peripheral blood CD4 and CD8 T-cell counts were determined using a Cyto-Stat tetraCHROME device (including labelling with CD45, CD3, CD4, and CD8) and acquisition on an FC500 flow cytometer (both from Beckman Coulter, Villepinte, France). Counts of naïve, memory, and terminal effector T-cells were determined as follows: CD8 and CD4 naïve T-cells were defined as CD45RA+CD62L+, peripheral memory T-cells were defined as CD45RA−CD62L−, and terminal effector CD8 T-cells were defined as CD45RA+CD62L−CD28−. The anti-CD62L antibody was obtained from BD Pharmingen (BD Biosciences, San Jose, CA, USA) and the CD4, CD8, and CD28 antibodies (Caltag) came from Life Technologies (Carlsbad, CA, USA).
3
Serum IgG and IgG4 Levels in IgG4-RD
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Blood samples were collected from both IgG4-RD patients and disease controls before steroids or other immunosuppressive medications were initiated. Total serum IgG and subclasses (IgG1 and IgG4) were measured by nephelometry (BNII analyser, Siemens, Surry, UK). An elevated serum IgG and IgG4 were defined as ≥16 g/l and ≥1.4 g/l respectively, the latter according to the Mayo HISORt criteria for AIP and local policy.
4
Serum Immunoglobulin Measurement in Clinical Research
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Total serum IgG and subclasses (IgG1, IgG2, IgG3 and IgG4) were measured by nephelometry using a BNII analyser (Siemens, Surry, UK) at the Clinical Immunology Department, Churchill Hospital. The normal range for serum immunoglobulin concentrations in the region was determined and validated by Oxford Immunology department, and is closely aligned with national immunoglobulin reference values; IgG 6-16g/l, IgG1 3.2-10.2g/l, IgG2 1.2-6.6g/l, IgG3 0.2-1.9g/l, IgG4 0.1-1.35g/l. For this study, an elevated serum IgG was ≥16g/l and serum IgG4 was ≥1.4g/l, as used in the Mayo HISORt criteria for AIP and in line with other studies. For patients with multiple serum immunoglobulin measurements, the first serum IgG4 level recorded was used. The prozone effect (falsely low serum IgG4 values) was accounted for using serial dilutions where necessary (19) .
5
Serum Biomarker Quantification Protocol
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Using stored samples, serum sST2, and sLR11 concentrations were measured by a colourimetric enzyme-linked immunosorbent assay (ELISA; sST2, Presage® ST2 Assay, Critical Diagnostics, San Diego, CA, USA; sLR11, MBS-167426, MyBioSource, San Diego, CA, USA). Inter- and intra-assay coefficients of variation were both < 10% for sST2, and < 15, and < 10%, respectively, for sLR11. hs-CRP concentrations were measured by a BNII analyser (Dade Behring, Newark, DE, USA) using a latex-enhanced immunonephelometric method.
6
Measurement of Serum Biomarkers in Fasting Subjects
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Fasting venous blood (10 mL) was collected from subject after fourteen hours of fasting period and kept in vacutainer tubes. Blood were divided into two heparinized and non heparinized tube. To separate serum, blood samples were immediately centrifuged. The samples were then stored at -80°C before analysis. Leptin concentrations were determined by the use of the ELISA Kit (eBioscience, USA) with inter- and intra-assay coefficient variances (CVs) of 6.7 and 8.9%, respectively and SOD was measured by the ELISA method using related kits (MyBioSource, USA) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively. The levels of hs-CRP were assessed by the turbidometric method (Pars azmoon, Tehran, Iran) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively. Plasma Hcy was assessed using high-performance liquid chromatography (HPLC) with colorimetric electrochemical detection with inter- and intra-assay CVs of 6.7 and 8.9%, respectively. TBARS levels were measured using the colorimetric technique as described by Uchiyama and Mihara (13 (link)) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively and SAA levels were measured using high-sensitivity method (ELISA) with a BNII analyser (Dade Behring) with previously validated techniques (14 (link)) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively.
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Fasting venous blood (10 mL) was collected from subject after fourteen hours of fasting period and kept in vacutainer tubes. Blood were divided into two heparinized and non heparinized tube. To separate serum, blood samples were immediately centrifuged. The samples were then stored at -80°C before analysis. Leptin concentrations were determined by the use of the ELISA Kit (eBioscience, USA) with inter- and intra-assay coefficient variances (CVs) of 6.7 and 8.9%, respectively and SOD was measured by the ELISA method using related kits (MyBioSource, USA) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively. The levels of hs-CRP were assessed by the turbidometric method (Pars azmoon, Tehran, Iran) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively. Plasma Hcy was assessed using high-performance liquid chromatography (HPLC) with colorimetric electrochemical detection with inter- and intra-assay CVs of 6.7 and 8.9%, respectively. TBARS levels were measured using the colorimetric technique as described by Uchiyama and Mihara (13 (link)) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively and SAA levels were measured using high-sensitivity method (ELISA) with a BNII analyser (Dade Behring) with previously validated techniques (14 (link)) with inter- and intra-assay CVs of 6.7 and 8.9%, respectively.
7
Baseline Evaluation of RBC Fatty Acids
8
Anthropometric and Metabolic Biomarkers
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Anthropometric measurements were taken after removing shoes, heavy clothing and belts. Body mass index (BMI) was calculated as weight [kg] divided by height2 [m2]. Blood pressure measurements were taken at the right arm after a rest period of at least five minutes in a sitting position and repeated three times at an interval of three minutes. The mean of the second and third measurement was calculated. A fasting venous blood sample was obtained from all study participants while sitting. Triglycerides were measured with the GPO-PAP-method (TGL Flex, Dade Behring, Marburg, Germany). Plasma high-sensitivity C-reactive protein (hsCRP) concentrations were measured using a latex enhanced nephelometric assay run on a BN II analyser (Dade Behring, Marburg, Germany). HsCRP values higher than 10 mg/l were excluded from the analysis to include only participants with subclinical inflammation. Glycated hemoglobin (HbA1c) was measured by high-performance liquid chromatography with spectrophotometric detection (Diamat Analyzer; Bio-Rad, Munich, Germany) and a coefficient of variation of 1.5%. The insulin resistance score (HOMA-IR) was calculated as fasting plasma glucose (mmol/l) × fasting serum insulin (mlU/l) / 22.5.
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