Ipsilateral brain was homogenized using Dounce Homogenizer in 10 volumes of NP40 cell lysis buffer (FNN0021, Thermo Fisher Scientific, USA) supplemented with 1mM phenylmethylsulphonyl fluoride (PMSF), and a protease inhibitor cocktail (Sigma-Aldrich). All steps were carried out at 4°C. The homogenate was centrifuged initially at 700 × g for 5 min to eliminate unruptured cells and debris, and then further centrifuged at 12,500g for 20 min and the supernatant was used to measure cytokine levels by ELISA. Tumor necrosis factor-alpha (TNF-α), IL-1β, interleukin-10 (IL-10), and IL-4 levels were measured by commercially available specific quantitative sandwich ELISA kits according to manufacturer instructions (eBioscience). The cytokine levels were normalized by total protein.
Specific quantitative sandwich elisa kits
Manufactured by Thermo Fisher Scientific
Specific quantitative sandwich ELISA kits are laboratory assays designed to accurately measure the concentration of a target analyte in a sample. The kits utilize a sandwich immunoassay format, where the analyte is captured between two antibodies and quantified through a color or fluorescent signal. These kits provide a reliable and standardized method for the quantitative detection of various biomolecules, enabling researchers to obtain precise measurements for their applications.
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2 protocols using specific quantitative sandwich elisa kits
1
Cytokine Quantification from Brain Homogenate
2
Cytokine Quantification from Brain Tissue
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Ipsilateral brain was homogenized using Dounce Homogenizer in 10 volumes of NP40 cell lysis buffer (FNN0021; Thermo Fisher Scientific) supplemented with 1 mm phenylmethylsulfonyl fluoride (PMSF) and a protease inhibitor cocktail (Sigma‐Aldrich). All steps were carried out at 4 °C. The homogenate was centrifuged initially at 700 g for 5 min to eliminate unruptured cells and debris and then further centrifuged at 12 500 g for 20 min, and the supernatant was used to measure cytokine levels by ELISA. Tumor necrosis factor‐alpha (TNF‐α), IL‐1β, interleukin‐10 (IL‐10) and IL‐4 levels were measured by commercially available specific quantitative sandwich ELISA kits according to the manufacturer's instructions (eBioscience). The cytokine levels were normalized by total protein.
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