Instrumentation for LC-MS/MS in Proteomics - LC-MS/MS in Proteomics: Methods and Applications - page 52

Fig. 3.2. Different operating modes of a triple quadrupole mass spectrometer. ( a )For
measurement of intact components, both quadrupoles Q1 and Q2 transmit all ions and
Q3 is operated in a scanning mode where only ions of a given m/z pass through the
chamber at a given time point during the scan. ( b ) For measurement of fragmentation
spectra of a component, Q1 is operated in a mass selective mode to transmit only ions
of the m/zof the desired precursor, Q2 is used as a collision cell, and Q3 is operated in a
scanning mode to measure the fragment ion m/z.( c ) For selective reaction monitoring,
Q1 transmits only the m/z of the precursor ion, Q2 is used as a collision cell, and Q3
measures only one fragment m/z.
quadrupole is operated in a mass selection mode to isolate a single
m / z component for subsequent analysis. The second quadrupole
is used as a collision chamber where applied energy and introduc-
tion of low levels of a collision gas (often nitrogen, but sometimes
helium or air) cause collisions and form fragments that can then
be measured using the third quadrupole ( Fig. 3.2b ) .
Quadrupoles are low-resolution and low mass accuracy ana-
lyzers. In a mass selection mode, they exhibit high sensitivity, but
in a scanning mode, they are insensitive compared to other analyz-
ers. Hence, triple quadrupoles are generally not the instruments
of choice for discovery of peptide identifications through frag-
mentation analysis. However, they are powerful in studies where
you already know how a component fragments and you want to
either detect this component at very low levels and/or quantify
its level. In experiments known as selected reaction monitoring
(SRM), a single mass can be monitored in the first quadrupole,
components are fragmented, and then the mass of a single frag-
ment ion is monitored in the final quadrupole ( Fig. 3.2c ) . By
monitoring a single reaction (given mass precursor forms a given
mass fragment), components can be detected at high sensitivity
and specificity, even in the background of a very complex mix-
ture. It is possible to cycle between a series of these SRM exper-
iments within an LC-MS analysis (a technique often known as
multiple reaction monitoring or MRM), allowing high-sensitivity