Microsoft excel based analysis software tools

Platelet poor plasma (PPP) and ePRP were assayed for the presence of human growth factors using a cytokine array (Cat. AAH-GF-1, RayBiotech, Peachtree Corners, GA, USA) and performed according to the manufacturer’s instructions. The data analysis has been employed by RayBiotech Microsoft^® Excel-based Analysis Software Tools for automated analysis (N = 40 individual human growth factor cytokines were detected).

Culture supernatants were collected at 24 h from duplicate wells seeded at 50,000 cells/well in a 24-well plate, with 0.5 mL total medium volume per well. Controls were prepared from the wells receiving complete medium only, without EVs. RayBio^® C-Series Canine Cytokine Array Kit 1, which can detect a total of 40 individual canine cytokines, was purchased from RayBiotech (RayBiotech, Peachtree Corners, GA, USA) and utilized according to manufacturer instructions. Briefly, after blocking, samples were placed in each well overnight at 4℃. The next day, samples were twice washed, and HRP-streptavidin incubation was performed overnight at 4℃ before a third wash. The next day, chemiluminescence detection was used to recognize the antibody levels expressed in the membrane. For comparison of their expression levels, internal positive controls were used to define the signal strength. Two duplicate spots were used for all samples. Data were analyzed in Microsoft® Excel-based Analysis Software Tools (RayBiotech) for each array kit via automatic analysis. The canine IFN-r (R&D Systems), IL-21, TNF- $\alpha ,$ and Rantes quantitative ELISA kits (RayBiotech) were applied to analyze these concentrations in the supernatant under the exposure with or w/o EVs.

Culture supernatants were collected at 24 h from duplicate wells seeded at 50,000 cells/well in a 24-well plate, with 0.5 mL total medium volume per well. Controls were prepared from the wells receiving complete medium only, without EVs. RayBio^® C-Series Canine Cytokine Array Kit 1, which can detect a total of 40 individual canine cytokines, was purchased from RayBiotech (RayBiotech, Peachtree Corners, GA, USA) and utilized according to manufacturer instructions. Briefly, after blocking, samples were placed in each well overnight at 4℃. The next day, samples were twice washed, and HRP-streptavidin incubation was performed overnight at 4℃ before a third wash. The next day, chemiluminescence detection was used to recognize the antibody levels expressed in the membrane. For comparison of their expression levels, internal positive controls were used to define the signal strength. Two duplicate spots were used for all samples. Data were analyzed in Microsoft® Excel-based Analysis Software Tools (RayBiotech) for each array kit via automatic analysis. The canine IFN-r (R&D Systems), IL-21, TNF- $\alpha ,$ and Rantes quantitative ELISA kits (RayBiotech) were applied to analyze these concentrations in the supernatant under the exposure with or w/o EVs.