The invention provides an enterovirus triple real-time fluorescent quantitative RT-PCR detection kit, which comprises a quantitative RT-PCR reaction solution, a standard substance and a reference substance, wherein, the quantitative RT-PCR reaction solution comprises a RT-PCR buffer solution, MgCl2, dNTPs, an EV universal primer, an EV universal fluorescent probe, an EV71 type fluorescent probe, a CA16 type fluorescent probe, heat resistant DNA polymerase and reverse transcriptase. The kit of the invention enables detection and genotyping to enterovirus universal type, enterovirus 71 type andcoxsackie virus group A 16 type by a one-step method. Compared with the current single detection kit, the kit of the invention is capable of saving the production cost and detection cost, raising theefficiency and time of the detection. Under the condition of ensuring no competition of multiple primers, the purpose of multi-channel detection can be achieved, so that the unification of the efficiency and technology is achieved.