Figures
Figure 1 : Characterization of proteins using denaturing CZE
conditions and denaturing ESI-MS as well as native CZE, native CIEF and
ACE coupled to native MS, respectively.
Figure 2 : Representation of the migration of cations in a CE-MS
setup depending on capillary coating, the resulting EOF and for the
required polarity. (A) shows the migration in an uncoated, (B) in a
neutrally coated capillary and (C) using a cationic (SMIL) capillary
coating. The length of the arrows represents the strength of the EOF
(green), the magnitude of analyte mobility (orange) and the magnitude of
the resulting apparent mobility (blue).
Figure 3. Evaluation of BGE composition on separation
performance of native CZE-MS. (A) Base peak electropherograms for
SigmaMab with 10 mM AmAc (AA), 10 mM AmF (AF), and 50 mM AmAc as the
BGE. The peaks labeled with black boxes in the electropherograms
represent the same mAb species. (B) Extracted ion electropherograms
(EIEs) of four different model proteins separated using 20, 40, and 60
mM AmAc as BGE, respectively. Separation improved with increasing AmAc
concentration. EIEs for the proteins were created using the three
highest charge states, respectively. Adapted and reprinted with
permission from ref (A) and (B). Copyright © 2021, Elsevier.
Figure 4. (A) Base peak electropherogram of E. coliribosomal proteins with 500 μM magnesium acetate in the BGE. (B)
Corresponding mass spectra of the dominant species in (A) collected at
(1) 53.72 and (2) 65.41 min allowed these species to be identified as
the 30S and 50S subunits. Base peak electropherograms of subcomplexes
and single proteins after rRNA removal with (C) no
Mg2+ and (D) 100 μM Mg2+ in the BGE.
Reprinted with permission from ref . Copyright © 2020 ACS, American
Chemical Society.
Figure 5. EIEs of a sample containing H2BK120ub and
H3K4,9,14,18ac analyzed with (a) standard (5 psi supplemental pressure)
and (b) high-resolution (2.2 psi supplemental pressure) CZE methods.
c−f: corresponding mass spectra generated by averaging the EIE peaks.
Reprinted with permission from ref . Copyright © 2021, American Chemical
Society.
Figure 6. Example for an ACE-nMS application showing a sheathless CE−MS
separation obtained for a H2O2-stressed
(A) NGmAb or (B) AAA-mAb sample using a BGE without FcRn (upper panel)
and containing FcRn (lower panel). The blue trace corresponds to the
extracted ion electropherograms (EIEs) of the double-oxidized /two red
stars), the green trace corresponds to the EIEs of mono-oxidized (one
red star), and the brown trace shows the EIEs of the non-oxidized mAbs
present in the mixture. Signal intensities are normalized. Reprinted
with permission from . Copyright © 2021 The Authors. Published by
American Chemical Society. Licenced under CC BY-NC-ND 4.0.
URL: https://pubs.acs.org/doi/10.1021/acs.analchem.1c03560.
Figure 7. Example for a nCIEF-nMS application. (A) Base peak
electropherogram of streptavidin (SA) analyzed by nCIEF-MS; (B)
deconvoluted mass spectra of seven SA charge variants separated from
nCIEF-MS. The inserted figure shows the zoomed-in EIEs for the
overlapping variants. Adapted and reprinted with permission from .
Copyright © 2022, American Chemical Society.
Table 1: Analysis of proteins using native CZE-MS, including MCE-MS and
ACE-MS.