Mouse cytokine array qam inf 1

Serum samples were screened in duplicates using a Mouse Cytokine Array QAM-INF-1 (RayBiotech, Atlanta, GA, USA), which consisted of slides coated with 40 different cytokines. The screening protocol was conducted following the manufacturer's guidelines, with minor adjustments as previously described [14 ]. In summary, the arrays were initially subjected to a blocking step, followed by an overnight incubation with 100 μL of conditioned medium. Subsequently, biotin-conjugated antibodies (1/250) were applied and incubated for 2 h, followed by an additional incubation with horseradish peroxidase (HRP)-linked secondary antibodies (1/1000) for 1 h. The membranes were incubated with a peroxidase substrate, and the outcomes were documented by the absorbance of the fluorescence. Quantitative analysis of the array data was conducted utilizing Array Vision Evaluation 8.0 (GE Healthcare Life Science, Marlborough, MA, USA).