Serum was collected by centrifugation at 2000 g for 5 min at 4°C, aliquoted, and stored at -70°C until analysis. A multiplex cytokine assay (M60009RDPD Bio-Rad, Hercules, CA, USA) was performed in accordance with the manufacturer's instructions using the Luminex 200 system instrument (Luminex Corporation).
Luminex 200 instrument system
Manufactured by DiaSorin
Sourced in United States
The Luminex 200™ Instrument System is a multiplex analysis platform that utilizes magnetic bead-based technology to perform simultaneous detection and quantification of multiple analytes in a single sample. The system is capable of conducting various assays, including immunoassays and nucleic acid-based tests.
Automatically generated - may contain errors
Lab products found in correlation
M60 009rdpd, by Bio-Rad (1 mentions)
Procartaplex, by DiaSorin (1 mentions)
Procartaplex, by Thermo Fisher Scientific (1 mentions)
Bio plex pro human cytokine 27 plex panel, by Bio-Rad (1 mentions)
Bobsoft software, by PerkinElmer (1 mentions)
Nanodrop one spectrophotometer, by Thermo Fisher Scientific (1 mentions)
Stellux chemiluminescent rodent insulin elisa kit, by ALPCO (1 mentions)
Mouse leptin elisa kit, by Crystal Chem (1 mentions)
7 protocols using luminex 200 instrument system
1
Serum Cytokine Profiling Procedure
2
Multiplex Cytokine Quantification
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We simultaneously analyzed a selection of 7 cytokines (BDNF, interleukin (IL)-10, IL-8, IL-6, leukemia inhibitory factor (LIF), interleukin 1 receptor antagonist (IL-1ra), and tumor necrosis factor alpha (TNF-α)), using a multiplex immunoassay kit (ProcartaPlex; Thermo Fisher Scientific, Waltham, USA). ProcartaPlex immunoassays are based on the principles of a ELISA, using 2 highly specific antibodies binding to different epitopes of 1 protein to quantitate all protein targets simultaneously using a Luminex 200 ™ System instrument (Luminex Corp., Austin, USA). The assays were performed according to the manufacturer's instructions. The standard curve was based on fiveparameter nonlinear regression (Fig. 1). Each cytokine concentration was then calculated by the curve.
3
Quantifying Alzheimer's Disease Biomarkers
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The amount of Amyloid Beta 40 (Aβ40), Amyloid Beta 42 (Aβ42), phosphorylated tau (pT181) and Total Human Tau was quantified according to manufacturer’s protocol using the bead-based Human ProcartaPlex™ kits from Thermo Fisher Scientific (Cat#EPX010-12,350–901, Cat#EPX010-12,351–901, Cat#EXP010-12,349–901, Cat#EXP010-12,348–901). A standard serial dilution and samples were prepared in duplicates to be read on Luminex 200™ Instrument System (Luminex Corporation, Austin TX). The concentration of each AD analyte was normalized by the total protein concentration for each mouse, and the normalized values were used for statistical analysis.
4
Multiplex Cytokine Profiling of HUVEC Secretome
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Culture medium of HUVECs was collected and analyzed with the Bio-Plex Pro™ Human Cytokine 27-plex Panel (Bio-Rad Laboratories, Hercules, CA, USA) in accordance with the manufacturer’s protocols. Briefly, culture medium was sequentially mixed with fluorescence-labeled capture beads and highly specific monoclonal antibodies. The mixture was then resuspended and irradiated. Concentrations of 27 cytokines and chemokines in the mixture were detected using the Luminex™ 200™ instrument system (Luminex, Austin, TX, USA). Data were analyzed with the xPONENT® software.
5
PNBoBs Assay for Prenatal Aneuploidy
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DNA was extracted from approximately 10 ml of amniotic fluid or 0.2 ml of umbilical cord blood using the QIAamp DNA Mini Kit (cat. no. 51306; Qiagen, Inc.), according to the manufacturer’s protocol. DNA concentration and purity were assessed using a NanoDrop One spectrophotometer (Thermo Fisher Scientific, Inc.,). The PNBoBs™ (cat. no. 3100-0020; PerkinElmer, Inc.) assay was performed according to the manufacturer’s instructions, which state that the assay can detect aneuploidies in chromosomes 13, 18, 21, X, and Y, as well as nine common microdeletion syndromes. The fluorescence of sample DNA bound to the beads was measured using a Luminex® 200™ instrument system (Luminex Corporation). Raw data generated by the instrument were analyzed using BoBsoft™ software (version 1.1; PerkinElmer, Inc.). Results were interpreted using the optimization method described in our previous report (Jiang et al., 2021 (link)).
6
Assessing Circulating Hormones in Mice
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Circulating levels of leptin and insulin were assessed by analyzing serum samples using ELISA kits: Mouse Leptin ELISA Kit (Crystal Chem, Elk Grove Village, IL, USA; 90030) and STELLUX Chemiluminescent Rodent Insulin ELISA Kit (Alpco, Salem, NH, USA; 80-INSMR-CH01). Pituitary hormone levels were measured in serum samples using the Mouse Pituitary Magnetic Bead Panel Milliplex Assay (EMD Millipore, Burlington, MA, USA; MPTMAG-49K), performed with the Luminex 200 Instrument System.
7
Quantification of Clazakizumab and Cytokine Levels
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Peripheral whole blood samples were collected in VACUETTE No Additive serum tubes (Greiner Bio-One) and centrifuged at 1890 × g for 10 min. The supernatant was collected in cryotubes and stored at -80 °C until measurement. Clazakizumab levels were quantified by enzyme-linked immunosorbent assay at ICON Laboratory Services, Inc‚ (Whitesboro, NY). Free (unbound to drug) and total (bound and unbound to drug) IL-6 were measured in 1:20 and 1:10 diluted serum samples, respectively, at Myriad Rules-Based Medicine (Austin, TX) using the fully automated Simoa HD-1/HD-X system (Quanterix; Billerica, MA). Soluble IL-6R (sIL-6R) was measured in 1:20 diluted serum samples using a commercial microbead assay (Human Magnetic Luminex Assay kit; Bio-Techne Ltd, Abingdon, United Kingdom). Measurements were performed on a Luminex 200 instrument system (Luminex Corp, Austin, TX). All samples were run in duplicates.
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