Effervescent MN patches were fabricated using polydimethylsiloxane (PDMS) (Dow Corning, Midland, MI) molds. Each MN was conical with a base radius of 150 μm, a height of 600 μm, and a tip radius of ~10 μm. The MNs were arranged in a 10 × 10 array in an area of 7 mm by 7 mm with a center-to-center spacing of 600 μm. To elevate the MNs, the patch backing contained an array of pedestals (base diameter, 600 μm; top diameter, 300 μm; and height, 350 μm) positioned at the base of each MN.

To fabricate MN patches, two solutions were cast sequentially onto the mold. The first casting solution contained 10% (w/w) solids, PLGA/LNG (60/40%, w/w) in diglyme/water (95%/5%, w/w). To prepare the first casting solution, 0.6 g of PLGA (50/50 lactide/glycolide molar ratio; inherent viscosity, 0.59 dl/g; Durect, Birmingham, AL) and 0.4 g of LNG (Chemo Industriale Chimica S.R.L, Saronno, Italy) were dissolved in a mixture of 7 g of diglyme (Sigma-Aldrich, St. Louis, MO), and 2.5 g of tetrahydrofuran (Thermo Fisher Scientific, Waltham, MA), and then LNG was crystalized and precipitated by slow evaporation of the tetrahydrofuran. Last, additional diglyme and deionized (DI) water were added to obtain the final casting solution. To fabricate MN patches containing Nile red (Sigma-Aldrich), 20 mg of Nile red powder was added into the casting solution and mixed to homogeneity.

Seven microliters of casting solution were applied to the top of the MN mold and then centrifuged at 3200g for 20 min to fill the mold. After waiting for 5 min, 20 μl of diglyme/water (95%/5%, w/w) was pipetted on the top of the mold, followed by centrifugation at 3200g for 20 min to wash residual casting solution on the top of the mold into the mold cavities. After that, the mold was placed in a 60°C oven with vacuum for 12 hours for drying.

Next, a second casting solution was prepared, consisting of 13% (w/v) PVP with two molecular weights (360/55 kDa, 50/50%, w/w; Sigma-Aldrich), and 4% (w/v) citric acid (Sigma-Aldrich) in pure ethanol (Koptec, King of Prussia, PA), with 5% (w/v) sodium bicarbonate (Sigma-Aldrich) suspended in the solution. Then, 80 μl of the second casting solution was gently applied to the dried PDMS mold surface to form the effervescent patch backing. For the control groups with noneffervescent backing, the second casting solution contained 13% (w/v) PVA (Sigma-Aldrich) and 13% (w/v) sucrose (Sigma-Aldrich) in DI water or 13% (w/v) PVP (360/55 kDa, 50/50%, w/w) in DI water. After drying in the chemical hood for 1 hour for the effervescent backing or 3 hours for the PVA/sucrose or PVP backing, the mold with effervescent backing or noneffervescent backing was placed in a desiccator overnight or for 2 days, respectively, at room temperature (20° to 25°C) for complete drying, after which a layer of adhesive paper was gently attached onto the top surface of the PDMS mold and used to carefully peel the patch from the mold, which was then stored in a desiccator until use.

To make MN patches for use in human participants, the first cast solution comprised 13% (w/v) PVA (Sigma-Aldrich) and 13% (w/v) sucrose (Sigma-Aldrich) in DI water. The second cast solution was the effervescent patch backing formulation containing PVP, citric acid, and sodium bicarbonate, as described above. The solutions were sterilized by passing through a sterile 0.2-μm filter (PES, VWR International, Radnor, PA). Molds were sterilized by autoclaving. Fabrication was carried out in a biological safety cabinet (Forma Class II, Thermo Fisher Scientific), and sample MN patches were tested to assure adherence to a low-bioburden specification.

Note: The content above has been extracted from a research article, so it may not display correctly.



Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.