Cytokine concentrations in cell-free supernatants and cell lysates were measured by commercial enzyme-linked immunosorbent (ELISA) kits, specific for: IL-17A (DY317 from R&D systems and 88-7176 from eBioscience), IL-17A/F (88-7117, eBioscience), IL-17B [ABKA2223 from Abnova (Taipei, Taiwan) and ab171344 from Abcam (Cambridge, United Kingdom)], IL-17F (887478, eBioscience), and CXCL8 (Mabtech, Nacka Strand, Sweden). ELISA detection limits were 4 pg/ml (eBioscience) and 15.6 pg/ml (R&D) for IL-17A, 30 pg/ml for IL-17A/F, 24 pg/ml (Abnova) and 10 pg/ml (Abcam) for IL-17B, 16 pg/ml for IL-17F, and 8 pg/ml for CXCL8.
Dy317
Manufactured by R&D Systems
Sourced in United States
The DY317 is a laboratory instrument used for the detection and quantification of protein and other biomolecules in samples. It utilizes a colorimetric detection method to measure the absorbance of light by the sample, which is proportional to the concentration of the target analyte.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using dy317
1
Cytokine Quantification in Cell Cultures
2
Quantifying IL-10 and IL-17A Cytokines
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Briefly, a 96-well plate (Eppendorf, Hamburg, Germany) was coated with monoclonal antibodies against IL-17A (DY317; R&D systems, Minneapolis, MN, USA), and IL-10 (DY217B; R&D systems) at 4°C overnight. After blocking with a phosphate-buffered saline (PBS)/1% bovine serum albumin/0.05% Tween 20 solution for 2 h at room temperature (22–25°C), the test samples and standards (recombinant IL-10 and IL-17A) were added to the 96-well plate and incubated at room temperature for another 2 h. The plates were washed four times with PBS and Tween 20 and then incubated with biotinylated human monoclonal antibodies against IL-10 and IL-17A for 2 h at room temperature. After washing, streptavidin-alkaline phosphatase-horseradish peroxidase conjugate (Sigma, St Louis, MA, USA) was added to the wells for 30 min, followed by incubation with 1 mg/mL p-nitrophenyl phosphate (Sigma, St Louis, MA, USA) dissolved in diethanolamine (Sigma, St Louis, MA, USA) to develop the color reaction. The reaction was stopped by the addition of 1 M NaOH, and the optical density of each well was measured at 405 nm.
3
Quantification of Cytokine Levels
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Briefly, a 96-well plate (Eppendorf, Hamburg, Germany) was coated with monoclonal Abs against TNF-α (DY210; R&D systems, Minneapolis, MN, USA), IL-17A (DY317; R&D systems), and IL-22 (MAB7822, 782-IL, BAM7821; R&D systems) at 4°C overnight. After blocking with PBS/1% bovine serum albumin/0.05% Tween 20 for 2 h at room temperature (22°C–25°C), the test samples and recombinant TNF-α, IL-17A, and IL-22 as standards were added to the 96-well plate and incubated at room temperature for another 2 h. The plates were washed 4 times with PBS and Tween 20, and then incubated with biotinylated human monoclonal Abs against TNF-α, IL-17A, and IL-22 for 2 h at room temperature. After washing, the streptavidin–alkaline phosphate–horseradish peroxidase conjugate (Sigma, St. Louis, MA, USA) was added to the wells for 30 min, and incubation with 1 mg/ml p-nitrophenyl phosphate (Sigma) dissolved in diethanolamine (Sigma) to develop the color reaction. The reaction was stopped by the addition of 1 M NaOH, and the optical density of each well was measured at 405 nm.
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