| Sourcefabric o.p.s. | Prague |
| Sova net | South Moravian |
| Spacemanic CZ, s.r.o. | |
| Spaneco spol. s r.o. | Prague |
| Speednet, s.r.o. | Ústí nad Labem |
| SPEL, a.s. | Central Bohemian |
| SPELEX, spol. s r.o. | South Moravian |
| Spendee, a.s. | |
| SpiNet, a.s. | |
| SportPass | |
| Springtide Ventures, s.r.o. | |
| Sprintel, s.r.o. | |
| Sprinter-studio, s.r.o. | |
| Sprinx | |
| SPSOFT, s.r.o. | |
| Stable Labs, s.r.o. | |
| Stable.cz | Liberec |
| Stadyum CRM | Prague |
| Stantum, s.r.o. | Prague |
| Star Lift | Prague |
| STARLIT, s.r.o. | Liberec |
| StartupBox z.s. | |
| StartupJobs | Prague |
| StartupJobs.com, s.r.o. | |
| StartUpStart | Prague |
| StartupYard | Prague |
| StickyPassword | South Moravian |
| Stofcom, s.r.o. | South Moravian |
| Stop Krádežím, spol. s r.o. | |
| Storm Type Foundry, s.r.o. | Prague |
| Storyous | |
| Storyous | |
| Strategic Consulting, s.r.o. | |
| Stratosyst, s.r.o. | |
| Stratox | |
| Stream Circle, a.s. | |
| Středisko společných činností AV ČR, v. v. i. | |
| Stren Tech, s.r.o. | |
| Studio 3P | |
| Studio 6.15 | |
| Studio dmm | South Bohemian |
| Studio DUHA, spol. s r.o. | |
| Studio Gabreta, spol. s r.o. | |
| Studolog | |
| Subreg | Prague |
| Suitcase Type Foundry | Prague |
| Suitest | |
| SUMA, spol. s r.o. | Prague |
| SUMO, s.r.o. | |
| Sun Marketing, s.r.o. | |