| Endora.cz | Liberec |
| Engeto, s.r.o. | South Moravian |
| Enter-Prise.net, s.r.o. | Prague |
| Envelon, s.r.o. | Prague |
| ENZON, s.r.o. | Prague |
| EpicAvatar | |
| Eqeus.com s.r.o. | South Moravian |
| ERA Components, s.r.o. | Prague |
| Eri Internet, s.r.o. | Hradec Králové |
| ESA BIC Czech Republic | Prague |
| Esense | Prague |
| ESPIRA Investments, s.r.o. | |
| Essence International, s.r.o. | South Moravian |
| eStarWeb.eu | Moravian-Silesian |
| eStation.cz | Hradec Králové |
| eStránky.cz | Prague |
| eStudio | Hradec Králové |
| ETARGET CZ, s.r.o. | Prague |
| Eternal, s.r.o. | Prague |
| Etheric Agency | |
| Etnetera | Prague |
| EuroPython 2023 | |
| Eurosoft - Control, s.r.o. | |
| EuroSoft, s.r.o. | |
| Eurosoftware, s.r.o. | |
| Eurosys, s.r.o. | |
| Eusoft, s.r.o. | |
| Evelup | |
| EVici webdesign, s.r.o. | Moravian-Silesian |
| Evocode, s.r.o. | |
| EVOTECH, s.r.o. | |
| ExaSoft Holding, a.s. | |
| Excalibur Army, spo. s r.o. | |
| EXPANDO global s.r.o. | Prague |
| Expo Data, spol. s r.o. | |
| fa Svoboda, s.r.o. | Prague |
| Fae.dev | |
| Fair Credit Czech, s.r.o. | |
| Fanatic Games Ltd | |
| Fast Communication, s.r.o. | Prague |
| Fastest Solution, s.r.o. | |
| FAYN Telecommunications, s.r.o. | South Moravian |
| FDLnet.CZ, s.r.o. | |
| FG Forrest, a.s. | |
| Finchainly | |
| Fine spol. s r.o. | Prague |
| FINEXT | |
| Fingood, s.r.o. | |
| Fintree | |
| First information systems, s.r.o. | Moravian-Silesian |