Delivery of type 1 diabetes antigen-specific immunotherapy using microneedles
The mass of PI coated on micrneedles (MNs) and the intradermal deposition of PI from PI-coated MNs into mouse skin were quantified by HPLC. The HPLC peaks relating to PI were integrated at a retention time of approximately 4.4 minutes. The chromatograms and peak areas are presented in .pdf files for each experiment. The peak areas have also been transferred and are presented in .xls files. Standards were analysed for each experiment and the concentration of unknown samples was interpolated from the appropriate standard curves (see .xls files). The standard curves were analysed by linear regression to generate an equation of the line (clearly stated in the .xls files) which was used to interpolate the unknown concentrations. These analyses were conducted in GraphPad Prism. PI mass was subsequently interpolated from the concentrations using equations clearly stated in the .xls files.
Microneedles were applied to mouse skin and the punctures were stained with methylene blue. Punctures were counted and the number of punctures and percentage puncture for each application was calculated and presented in Microsoft Excel (in the form of a .xls file). The equation used to calculate percentage puncture is shown in cell C15. Primary puncture in columns D and E refers to the entry points of the MNs into the skin. Secondary puncture in columns G and H refers to the exit of the MNs from the adjacent side of the skin fold. Average percentage puncture is shown in cells E9 and H9 and standard deviation of percentage puncture is shown in cells E10 and H10.
Images of the punctured skin were acquired using the Nikon D5000 12.3MP Digital SLR Camera. The image is in the form of a .jpg file.
Scanning electron microscopy (SEM) images of microneedles were acquired using the FEI XL-30 FEG environmental scanning electron microscope. In the image file names: C refers to uncoated microneedles; CC refers to microneedles coated in proinsulin; S refers to microneedles coated in proinsulin and applied to mouse skin; SC refers to uncoated microneedles applied to mouse skin. In the image file names, x76 and x250 refer to magnification. The images are in the form of .tif files.
Flow cytometry data is in the form of .fcs files. These were processed in FlowJo Software and this processed data is available in the form of .wsp files for each of the three separate experiments conducted. FlowJo software is required in order to access the .fcs and .wsp files. Processed data from FlowJo is presented in Microsoft Excel (in the form of a .xls file). Statistical analysis was conducted using Graphpad Prism.
3H-thymidine incorporation was used as a cell proliferation readout in in vitro studies. Raw 3H-thymidine incorporation readings (ccpm) are presented in a Microsoft Word document (.doc) and have been transferred to a Microsoft Excel file (.xls) for clarity and processing for each of the three separate experiments. Readings for the experimental samples are presented under “PI+CD8+DC” and readings for the control samples are presented under “PI+DC”. Stimulation index was calculated using the equation shown in cell B22. The stimulation index data was used to plot a graph in GraphPad Prism. Statistical analysis was conducted using GraphPad Prism.
Research results based upon these data are published at http://doi.org/10.1016/j.jconrel.2020.02.031