For all the assembly periods tested, the Belly Dancer® and the end-over-end mixer resulted in similar or higher average NPs areal density, compared to the vortex mixer (Figure 3a). For the former two mixing methods, as the assembly period increased from 30 min to 60 min, the count of NanoBEADS with attachment numbers higher than 10 NPs/µm2 cell area showed an appreciable increase while the number of bacteria without any NP attached (0 NPs/µm2) decreased (Figure 3c,d). Upon increasing the assembly time to 90 min, more NanoBEADS with attachment density lower than 10 NPs/µm2 were observed (Figure 3c,d), resulting in an overall decrease in average attachment density (Figure 3a). This reduction in attachment density can be attributable to the presence of residual culture media in the assembly volume, which led to some bacteria growth during the more extended assembly period. Therefore, 60 min was identified as the optimal assembly duration. B-800-60 and E-800-60 cases had significantly higher NPs attachment density than V-800-60 case (p<0.05). Additionally, there was a significant difference in the NPs attachment density in E-800-60 comparing to the other two durations (p<0.01). Moreover, the E-800-60 case had a statistically significant higher average NP attachment density than the NanoBEADS constructed using a vortex mixer, except for the V-100-60 case, where the difference was not statistically significant. Altogether, our results suggest that the Belly Dancer® mixer used with 800 µL assembly volume for 60 min (B-800-60), and the end-over-end mixer used with 800 µL assembly volume for 60 mins (E-800-60) were the optimal sets of assembly parameters, among the combinations tested.