3.4 Protein stability in electrolyte at sequential time frame
The major concern of selecting the bio-NPs as RM was its degradation and deactivation. To learn the stability of bio-NPs, the candidate proteins selected as the sub-10 nm RM, BSA and ubiquitin, which exhibited consistent results with literature [6,19,21], were suspended in 20 mM AmAc with the respectively protein concentrations of 0.1 mg/ml and 0.5 mg/ml. The size distribution and representative diameter of BSA and ubiquitin were monitored as a function of time to confirm the dispersive and structural stabilities of these proteins. In Table S2 , the diameters of BSA and ubiquitin measured on sequential days were recorded and the size variance of their representative diameters remained < 10 % difference from literature.
On the 63th day of BSA suspension, the detected diameter declined to 6.6 nm and had a 4.3 % difference from the value obtained on the 1st day. A new BSA solution at the sample protein concentration was prepared and tested on the same day to approve the change in size, which contributed to the structural alternation of BSA in electrolyte. Figure S3 indicated the similar size distributions and the same measured representative diameter of BSA generated from both the 63th-day BSA solution and the fresh BSA solution measured on the same day. In association with the diameter of the 1st-day BSA solution, the shift in size distribution was evident, from which not only the change in monomer but also dimer. On the 120th day of BSA suspension, the representative diameter of BSA was again measured as 6.9 nm, which suggested the slight change in diameter was the floating of the ES-SMPS system. As a result, the consistent diameters of BSA and ubiquitin obtained in this experiment and reported in literature illustrated the repeatability of BSA and ubiquitin and presented their suitability as the sub-10 nm and sub-5 nm RM.