Native mass spectrometry is widely used to determine the stoichiometries and binding constants of noncovalent interactions in solution. One challenge is that multiple analytes in a single electrospray droplet can aggregate during solvent evaporation, which will bias the distribution of oligomeric states observed during gas-phase measurements. Here, measurements of solution flow rates, electrospray currents, droplet size distributions, and nonspecific aggregation are used in conjunction with Poisson statistics to characterize the factors that control nonspecific aggregation during typical native mass spectrometry experiments. Using electrokinetic nanoelectrospray ionization and a 30 nA current, low flow rates of less than 10 nL min−1 and initial droplets with mean diameters of ∼60 nm were observed. For solutions containing 4 μM analyte under these conditions, Poisson statistics and charge-reduction drift tube ion mobility spectrometry both indicate that ∼90% of the desolvated, occupied droplets contain a single analyte. Initial droplet sizes and contributions from nonspecific aggregates both increase with increasing current. Ion mobility mass spectrometry analysis of the ions produced using these conditions without charge reduction exhibit even less nonspecific aggregation (∼2%). All approaches indicate that increasing the ionization current increases the flow rate, droplet size distribution, and extent of nonspecific aggregation. These results provide detailed insights into the role of small initial droplets in the success of native mass spectrometry.
|Original language||English (US)|
|Number of pages||8|
|Journal||International Journal of Mass Spectrometry|
|State||Published - Sep 2017|
Bibliographical noteFunding Information:
M.F.B. and K.L.D. acknowledge support from the University of Washington. Support from US National Science Foundation Grant CBET- 1133285 was used to the purchase of consumables for operation of the charge reduction ESI experiments. D.R.O. acknowledges support from a National Science Foundation Graduate Research Fellowship.
© 2016 Elsevier B.V.
- Charge reduction
- Noncovalent interactions
- Nonspecific interactions
- Protein complexes