Biology Reference
In-Depth Information
until it becomes unstable upon reaching its Rayleigh limit. At this
point, the droplet deforms and emits charged jets in a process
known as Coulomb fission. Repeated charge-induced droplet dis-
integrations lead ultimately to small highly charged droplets capa-
ble of producing gas-phase ions. Multiply charged ions such as
[M +
n
H]
n
+
are often observed. Polypeptides tend to be variably
protonated at arginine, lysine, and histidine residues. Multiple
charging enables mass analyzers with limited
m
/
z
ranges to ana-
lyze higher molecular mass molecules. In addition, ESI allows non-
covalent protein complexes to be ionized intact, showing its utility
for determining the quaternary structure of proteins [
42
].
A shortcoming of ESI is its susceptibility to ion suppression at rela-
tively low salt concentrations, > ~1 mM, which means that biologi-
cal samples need to be desalted before ESI-MS.
Several theories have been proposed to explain the final pro-
duction of gas-phase ions: the Charge Residue Model (CRM)
[
43
]; the Ion Evaporation Model (IEM) [
44
,
45
]; and a model
invoking combined charged residue-field emission [
46
]. Whatever
mechanism operates, and because ion formation involves extensive
solvent evaporation, typical solvents for electrospray ionization are
mixtures of water with volatile organic solvents (e.g. methanol,
acetonitrile). These solvents are typically employed in reverse-
phase high-performance liquid chromatography (RP-HPLC). Due
to the ease of coupling LC directly to the ion source, the combina-
tion of liquid chromatography and mass spectrometry has entered
the realm of routine analysis.
The physics behind MALDI is also debated [
47
-
49
]. Ionization
is triggered by firing a pulsed laser beam at a dried-droplet spot of
co-crystallized matrix and analyte molecules usually placed in a
vacuum chamber. The matrix (in large excess) absorbs the laser
energy and is desorbed and protonated. The ionized matrix trans-
fers the proton to analyte molecules thus generating M + H
+
ions.
Since the typical MALDI source generates pulse of ions, this ion-
ization method is most compatible with spectrometers which function
as pulsed-ion detectors (TOF) and with instruments which trap
the ions for later analysis (ion traps and FT-ICR instruments).
Mass analyzers which operate on a continuous beam of ions, such
as magnetic sector and quadrupole instruments, are generally not
suitable for a pulsed-ion source. Although MALDI sources that
operate at atmospheric pressure (AP) have been developed, which
approach the sensitivity of vacuum MALDI [
50
], still the most
natural form of interfacing a liquid chromatograph to MALDI-MS
is off-line. However, it is now well established that time-of-fight
mass spectrometry involving instruments with independent
(orthogonal) axes for ion generation and mass analysis, generally
referred to as orthogonal acceleration time-of-flight (oa-TOF)
mass spectrometry, are well suited to continuous ion sources [
51
].
This approach allows synchronization of ion formation with mass
