Serum levels of IL-6, TGF-α, and TNF-α were determined using premixed multiplex human magnetic Luminex assays (R&D, #LXSAHM-11) according to the manufacturer’s instructions. Data acquisition was performed using the Bio-Plex MAGPIX Multiplex reader (BioRad Laboratories, Hercules, CA, USA), which uses Luminex fluorescent-bead-based technology (Luminex, Austin, TX, USA). Serum levels of TGFβ1 (EBioscience, San Diego, CA, USA, #BMS249/4) and IL-15 (Thermo Scientific, Waltham, MA, USA, #BMS2106) were evaluated with enzyme-linked immunosorbent assay (ELISA, Ratavartijankatu, Finland). The optical density was measured spectrophotometrically at a wavelength of 450 nm according to the manufacturer’s instructions. All analyses were performed in duplicate. A 4-PL standard curve was created using MyCurveFit Beta Software (https://mycurvefit.com/ , accessed on 11 April 2023).
Lxsahm 11
Manufactured by R&D Systems
The LXSAHM-11 is a laboratory instrument designed for general-purpose use. It features adjustable speed and temperature controls. The core function of this product is to facilitate sample preparation and processing tasks in research and laboratory settings.
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Lab products found in correlation
4 protocols using lxsahm 11
1
Serum Cytokine Profiling Protocol
2
Plasma Cytokine Measurement
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To measure systematic inflammation levels, plasma cytokines including MCP-1, IL-6, IL-10 and IL-12 were measured using a Human Magnetic Luminex Screening Assay (11 PLEX) (R&D LXSAHM-11). More details are given in the Supplementary Materials and Methods
3
Multiplex Profiling of Splenic Soluble Factors
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The spleen samples were collected and stored at −80 °C. The soluble factors in the spleen were quantified on the Luminex FLEXMAP 3D multiplexing platform. A mouse premixed multi-analyte kit (LXSAHM-11; R&D Systems, Shanghai, China) was used to measure the concentrations of a panel of soluble factors, including interleukin-1 alpha (IL-1α), IL-2, IL-4, IL-10, IL-17 A, C–C motif chemokine ligand 2 (CCL2), CCL3, CCL4, interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), and granzyme B (GZMB) in one sample at the same time. Luminex assays were performed as the manufacturer's protocol suggested, and each group contains 6 biological replicates. The samples were read on a Luminex FLEXMAP 3D System. The raw data were processed with the Milliplex Analyte program by using a five-parameter logistic regression analysis.
4
Multiplexed Biomarker Profiling of pEVs
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The pEVs’ protein content of BDNF, APP, NSE, NPTX‐2, α‐Syn, DJ‐1, MMP‐9, S100B, PrGN, LCN‐2, and ANGPTL‐4 was determined by the multiplex Luminex assay (LXSAHM‐11, R&D Systems, Inc.,). Similarly, the accumulation of the protein fragments Aβ1‐40, Aβ1‐42 and different forms of the protein tau (t‐tau and p‐tau181) were evaluated by another Luminex assay (HNABTMAG‐68K, Millipore‐Sigma). Data were processed using an analytical software coupled to the Luminex 100/200 machine (Xponent 4.2, USA) and results were normalized according to the total pEVs’ protein amount. Some values were below the limit detection, which reduced the average number of participants per group but maintained adequate statistical analysis. Limit detection sensibility for the analytes is described (Table S2).
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