Suprachoroidal injection is an emerging technique for drug delivery to the posterior segment, which is hard to reach by non-invasive approaches. However, the injection technique varies and the associated ocular safety is not well understood. In addition, it is not clear if drug formulation is a major factor in optimizing pharmacodynamics using this technique. The current study was designed to compare the suprachoroidal injection of different drug formulations and to characterize the safety and pharmacodynamics of triamcinolone acetonide (TA) delivered by this technique. Both indocyanine green (ICG) solution and TA suspension, at 50μL, 100μL, and 150μL, were suprachoroidally injected and intraocular pressure (IOP) tonometry, fundus photography, and electroretinography were performed over multiple time points up to eight weeks. After 50μL TA (Kenalog-40) suprachoroidal injection, 4–5 animals at 7 time points were sacrificed for aqueous, vitreous, retina, and plasma collections. TA was quantitated using ultra-performance liquid chromatography tandem mass spectrometry. For comparative efficacy study, 50μL (2mg) suprachoroidal TA versus 20mg subtenon TA were performed 4weeks before induction of experimental uveitis with 10ng of intravitreal lipopolysaccharide. After suprachoroidal injection, IOP had an acute elevation, higher volume caused higher IOP (p<0.0001). Equivalent volume of ICG solution led to a significantly smaller IOP elevation than after TA suprachoroidal injection. This finding suggests better distribution of ICG solution than TA suspension in the suprachoroidal space. Following a 50μL suprachoroidal injection, peak TA concentration in the aqueous was below 1ng/mL. In contrast, the posterior vitreous and retina had 1912ng/mL and 400,369ng/mL TA, respectively. Maximum TA in plasma was 11.6ng/mL. Drug exposure to the posterior retina was 523,910 times more than that to the aqueous and 29,516 times more than systemic TA exposure. In the treatment of lipopolysaccharide-induced uveitis, compared with 20mg subtenon injection, suprachoroidal 2mg TA demonstrated much better efficacy with significantly less aqueous humor cells and lower vitreous opacity scores (p<0.05). Histology showed much less vitreous inflammation in the suprachoroidal injection group (p<0.0001). It seems that a 50μL suprachoroidal injection of TA was well tolerated in rabbit eyes and demonstrated excellent penetration into the posterior retina, providing better therapeutic effect than subtenon 20mg TA.