Since the discovery of the invasive soybean aphid, Aphis glycines Matsumura, in the USA in 2000, populations have expanded throughout the Midwestern USA and Canada. Its negative economic impact on soybean [Glycine max (L.) Merr.] yield provides the impetus to continue investigating the effectiveness of host plant resistance which has been affected by soybean aphid biotypes that overcome soybean plants containing Rag (resistance to A. glycines) genes. In this study, 14 aphid clones collected on soybean and buckthorn (Rhamnus cathartica L.) plants along with four known aphid biotypes (from our stock collection) were evaluated in no-choice assays by quantifying the number of aphids on soybean genotypes with Rag genes and the susceptible cultivar Williams 82. No-choice assays were conducted using detached leaves and whole plants. Inconsistent responses of the biotypes from the stock collection by using detached leaves confirm the efficacy of whole plant assays to accurately differentiate biotypes. None of the biotypes or field clones from Illinois, Indiana and South Dakota overcame the resistance of soybean genotypes LD14-8004 (Rag1) and PI437696. A soybean aphid clone from Wooster, Ohio readily colonized LD14-8002 (Rag2), but did not overcome any of the other resistance genes indicating that it is different from biotypes 3 and 4, which also overcome LD14-8002 (Rag2) and, respectively, LD14-8004 (Rag1) and LD14-8006 (Rag3).