Systematic assessment of microneedle injection into the mouse cornea

Forty eyes of 20 129？Sv/J mice were tested. India ink was intrastromally injected using 30° beveled 33？G needles, tri-surface 25° beveled 35？G needles, or hand-pulled and 25° beveled glass needles. Each eye received a single injection of a volume of 1 or 2 μL. Corneoscleral buttons were fixed and flat mounted for computer-assisted quantification of the affected corneal area. Histological assessment was performed to investigate the intrastromal location of the injected dye.A mean corneal area of 5.0 ±1.4？mm2 (mean？±？SD) and 7.7 ±1.4？mm2 was covered by intrastromal injections of 1 and 2 μL, respectively. The mean percentage of total corneal area reached ranged from 39% to 53% for 1 μL injections, and from 65% to 81% for 2 μL injections. Injections using the 33？G needles tended to provide the highest distribution area. Perforation rates were 8% for 30° beveled 33？G needles and 44% for tri-surface beveled 35？G needles. No perforation was observed with glass needle; however, intrastromal breakage of needle tips was noted in 25% of these cases.Intracorneal injection using a 30° beveled 33？G needle was safe and effective. The use of tri-surface beveled 35？G needles substantially increased the number of corneal perforations. Glass needles may break inside the corneal stroma. Injections of 1 μL and 2 μL resulted in an overall mean of 49% and 73% respectively of total corneal area involved.Effective corneal gene therapy will require safe and effective gene delivery. However, lacrimation, mucus, blinking of the eyelid, and tight junctions of the corneal epithelium are a significant obstacle to genetic vectors [1,2]. Accordingly, intrastromal or intracameral injections are generally used for delivery of genetic therapeutics to subepithelial corneal layers [1].Corneal thickness substantially varies among different animal species [3]. It is reported to be approximately 354？μm in rabbits and 170？μm in rats, while it is only 89.2 to 123.8？μm in mice [3,4]. Therefore, many studies i