Concentrations of 40 cytokines in the serum samples were measured with a commercial quantitative immuno-microarray (Quantibody Human Cytokine Array 1, RayBiotech, Inc., Norcross, GA, USA) according to the manufacturer’s instructions, and results were analyzed using the RayBiotech Q Analyzer program. Glass chips were washed after incubation in blocking buffer for 30 min, and each well was overlaid with 100 μL of diluted sample. After overnight incubation at 4°C and extensive washing, the chips were incubated for 1 h with detection antibody and then washed. AlexaFluor 555-conjugated streptavidin was then added and incubated for 0.5 h at room temperature. Signals (Cy3; 555 nm excitation, 655 nm emission) were scanned using a Genepix 4000B laser scanner (Axon Instruments, Foster City, CA, USA).
Quantibody human cytokine array 1
Manufactured by RayBiotech
Sourced in United States
The Quantibody Human Cytokine Array 1 is a multiplexed quantitative sandwich ELISA-based assay designed for the simultaneous measurement of multiple human cytokines in a single sample. The array contains antibodies specific to 40 different human cytokines, chemokines, and growth factors, allowing for the simultaneous quantification of these analytes in a single experiment.
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3 protocols using quantibody human cytokine array 1
1
Quantitative Cytokine Profiling in Serum
2
Quantifying Cytokine Secretion with ELISA
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IL-6 cytokine secreted by cells was quantified using LEGEND MAX Human IL-6 ELISA Kit (Biolegend #430507). Media supernatant collected from culture of various cells were incubated and stained with reagents on a pre-coated 96-well strip plate as per manufacturer’s instructions. A microplate reader (Enspire) was used to measure the absorbance at 450 nm.
Quantibody® Human Cytokine Array 1 (RayBiotech) is a multiplex ELISA system for quantitative measurement of multiple cytokines simultaneously. Sample preparation and analysis was performed according to the manufacturer’s instructions. Further statistical analysis was performed using GraphPad.
Quantibody® Human Cytokine Array 1 (RayBiotech) is a multiplex ELISA system for quantitative measurement of multiple cytokines simultaneously. Sample preparation and analysis was performed according to the manufacturer’s instructions. Further statistical analysis was performed using GraphPad.
3
Quantifying Cytokine Levels in Stimulated Blood
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Fresh blood (1 mL) was stimulated with TB-specific antigens (ESAT-6 and CFP-10) or PBS (negative control) for 20 h at 37°C. Then, centrifugation was performed at 4000 g, and the plasma was collected. A commercial quantitative immuno-microarray (Quantibody Human Cytokine Array 1, RayBiotech, Inc., Norcross, GA) was used to measure the concentrations of 40 cytokines in the plasma samples. The manufacturer protocol was followed, and the results were analyzed using the RayBiotech Q Analyzer program. Briefly, after 30 min of incubation with sample diluent, the glass chips were washed, and each well was overlaid with 100 μL diluted sample.
After overnight incubation at 4°C and extensive washing, the detection antibody was added for 1 h and then washed away. AlexaFluor 555-conjugated streptavidin was then added for 2 h at room temperature. The signals (Cy3; 555 nm excitation, 655 nm emission) were scanned and extracted using a Genepix 4000B laser scanner (Axon Instruments, Foster City, CA, USA). The total amounts and concentrations of the biomarkers were evaluated in this study. The concentrations, expressed in pg/mL, were calculated using RayBiotech Q Analyzer software against a standard curve determined for each biomarker. The total amount of each biomarker was determined in pg by multiplying the concentration by the sample volume.
After overnight incubation at 4°C and extensive washing, the detection antibody was added for 1 h and then washed away. AlexaFluor 555-conjugated streptavidin was then added for 2 h at room temperature. The signals (Cy3; 555 nm excitation, 655 nm emission) were scanned and extracted using a Genepix 4000B laser scanner (Axon Instruments, Foster City, CA, USA). The total amounts and concentrations of the biomarkers were evaluated in this study. The concentrations, expressed in pg/mL, were calculated using RayBiotech Q Analyzer software against a standard curve determined for each biomarker. The total amount of each biomarker was determined in pg by multiplying the concentration by the sample volume.
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