The US Environmental Protection Agency (EPA) conducted a Wadeable Stream Assessment (WSA) of all wadeable streams and rivers in the conterminous US between 1999 and 2005. The assessment was led by the EPA Office of Water, in cooperation with EPA regions, states, tribal nations, and the EPA Office of Research and Development (ORD). The WSA was implemented as 2 large-scale regional surveys of streams and rivers. Both studies used EPA's River Reach File (RF3) as the basis for the sample frame. The Environmental Monitoring and Assessment Program (EMAP) Western Pilot Study, conducted by ORD in cooperation with EPA Regions 8, 9, and 10 and 12 western states, assessed all streams and rivers in the 12 western states (EMAP-West). A stratified, unequal probability survey design (50 sites/state and additional sites in 5 intensive study areas) was used to select sites from all streams and river segments coded as perennial in RF3. The unequal selection depended on Strahler order, aggregated Omernik level III ecoregion, and special study region. The WSA study used the EMAP-West wadeable streams (WSA-West) and implemented a new design for the remaining 36 eastern conterminous states (WSA-East). The WSA-East design was an unequal probability survey design with unequal selection depending on Strahler order, Omernik Level II ecoregion, and EPA region. RF3 includes 5.29 million km of rivers and streams, of which 39% (2.07 million km) are coded as perennial. The WSA sample frame included 2.84 million km of streams (54% of the total length in RF3), of which 2.24 million km were in WSA-East and 0.60 million km were in WSA-West. Each selected site was classified on the basis of wadeability and the presence of flowing water. The estimated length of wadeable streams and rivers in the 48 conterminous states was 1.30 ± 0.025 (SE) million km (45.7 ± 1.1% of the stream length in the sample frame). Of this wadeable stream length, 78.6 ± 1.0% (1.02 million km) was estimated to be appropriate for sampling. Nationally, 11.5 ± 0.8% and 5.2 ± 0.6% of this length could not be sampled because of access denial or physical inaccessibility, respectively. The proportion of length affected by access denial was higher in Southern Plains, Northern Plains, and Xeric West aggregated ecoregions, whereas stream length affected by physical inaccessibility was greatest in the Western Mountains aggregated ecoregion. Improvements in the sample frame (RF3 and its successors National Hydrography Database [NHD] and NHD-Plus) would reduce field costs for national surveys.