Snow pool Aedes mosquitoes are identified easily and accurately by microscopic examination of 4th instars or male genitalia. Early instars and females cannot be identified accurately. We demonstrate that restriction fragment-length polymorphisms (RFLP) in the internal transcribed spacer (ITS) of the nuclear ribosomal DNA cistron are specific to each of 13 snow pool Aedes species found in northern Colorado. The ITS was amplified by the polymerase chain reaction (PCR) and digested with AluI and MspI restriction endonucleases. Differences in the sizes of digested fragments among the 13 species were so slight that they could only be resolved with polyacrylamide gel electrophoresis and visualized with silver staining. A key to species found in the Rocky Mountains of northern Colorado was constructed using the PCR-RFLP patterns. Three of the species were collected in California and had identical PCR-RFLP patterns, indicating that restriction sites in the ITS may be conserved within some species.