Aquatic macroinvertebrates are vital to stream ecosystem function, including as food sources for higher trophic levels and are used as indicators of stream condition due to sensitivities to water quality and habitat disturbance. This study aims to synthesize several aquatic macroinvertebrate bioassessment methodologies into a single protocol that better captures biodiversity and reproducibility across sites and time. Incorporating sampling protocols from U.S. Environmental Protection Agency (USEPA), Georgia Environmental Protection Division (GAEPD), and South Carolina Department of Environmental Services (SCDES), to create a revised sampling protocol with the standardization and habitat-based sampling approach of USEPA/GAEPD, while sampling large areas and multiple habitat types to increase captured biodiversity, found in the SCDES protocol. To determine the number of samples within a stream reach needed to increase the biodiversity sampled while remaining standardized, we conducted a preliminary study using four different streams at the Savannah River Site (Aiken, SC, USA) across a gradient of size and degradation. We took samples from six leaf packs, six snags, five bank samples, and nine sand samples (three from pools, runs, and flats each). Specimens were identified to the Family level. To understand how biodiversity varied with sample number and habitat type, species accumulation curves and Venn diagrams were created. Preliminary analysis showed that 80% of the Family level aquatic macroinvertebrate biodiversity was found within three samples for leaf packs, snags, and banks, and six samples for sand habitats (two pools, flats, and runs each). Our preliminary study found 37, 50, 55, and 62 families at our four sites, compared to 15, 25, 14, and 30 families respectively at the same sites found during a 2022 SCDES protocol sampling event. Leaf packs largely contained families found within other habitats: only 0-5.4% of families were unique to leaf packs. These results allowed us to reduce the number of samples taken in each habitat in our follow up sampling comparing this methodology, SCDES, and GAEPD in the same streams, while also revealing the biodiversity that can be missed with non-standardized, less intensive sampling efforts.