Insects with over 30,000 aquatic species are known as very successful arthropods in freshwater habitats. Some of them are applied as biological indicators for water quality control, as well as the main food supply for fishes and amphibians. The faunistic studies are the basic step in entomological researches; the current study was carried out emphasizing on the fauna of aquatic insects in Karaj River, northern Iran. A field study was carried out in six various sampling site of Karaj River during spring 2013. The aquatic insects were collected using several methods such as D-frame nets, dipping and direct search on river floor stones. Specimens were collected and preserved in Ethanol and identified by standard identification keys. Totally, 211 samples were collected belonging to three orders; Plecoptera, Trichoptera and Ephemeroptera. Seven genuses (Perla, Isoperla, Hydropsyche, Cheumatopsyche, Baetis, Heptagenia and Maccafferium) from five families (Perlidae, Perlodidae, Hydropsychidae, Batidae, Heptagenidae) were identified. The most predominant order was Plecoptera followed by Trichoptera. Karaj River is a main and important river, which provides almost all of water of Karaj dam. So, identification of aquatic species which exist in this river is vital and further studies about systematic and ecological investigations should be performed. Also, monitoring of aquatic biota by trained health personnel can be a critical step to describe water quality in this river. Understanding the fauna of aquatic insects will provide a clue for possible biological control of medically important aquatic insects such as Anopheles as the malaria vectors.