Squirrel Tutorial
Post – Pre-Process Steps
 Airbeam correction
 PToF DVa, DC markers
 Fragmentation checks
Donna Sueper
Aerodyne, University of Colorado, Boulder
Tutorial Outline
Squirrel Quick-Look Flow Chart
Squirrel Complete Analysis Flow
Chart
Calculation discussion, Screen Shots
Airbeam correction
PToF DVa, DC markers
Frag checks
HDF Index Tab
Squirrel Quick-Look Flow Chart
Get Index
Pre-Process
time series,
avg spectrum
PToF Tab
MS Tab
_p data (DAQ sticks)
image,
avg size dist,
size binned spectrum
Squirrel Analysis Flow Charts
baseline fitting params
Modify SI
Ion_user
Pre-Process
Airbeam
size dist,
mass spec,
image, etc.
corr_fact
Do Corrections
PToF
time series
avgs, etc.
Corrections Tab
size dist,
mass spec,
image, etc.
m/z fitting params
Check baseline
_p data (recalc sticks)
MS Tab
time series
avgs, etc.
PToF Tab
MS Tab
_p data (DAQ sticks)
Check m/z calibration
Frag Checks Tab
Pre-Process
Get Index
PToF Tab
Get Index
Complete Analysis
HDF Index Tab
HDF Index Tab
Quick-Look
_p data (dc marker)
Calc Loadings
Loading waves
Tweak
Frag table
Diagnostics
Plot
Complete Analysis > Corrections
Select the Corrections tab
There are 5 subtabs:
• Flow Rate
• Airbeam
• PToF
• Errors
• IE Cals
When the gold Do Corrections button is
pressed, the actions as checked in each
subtab will be performed. For example:
Airbeam
PToF – DVa
PToF – DC markers
Airbeam Correction Discussion
The airbeam value is a measure of the overall amount of sample we
measure. For each run the airbeam value (mass 28) changes, often by a
relatively small amount.
In the diagnostics graph,
we see the PToF and MS
airbeam. For the moment
we consider only the MS
airbeam.
We want to take an average of the airbeam at a specific region (airbeam
reference value runs) and compute the ratio of this value to the airbeam
values at all other runs.
Correction factor = (airbeam avg/airbeam).
Complete Analysis > Airbeam correction
On the Corrections tab, select the Airbeam (AB) tab
For this example we will use the Auto-set option.
Select airbeam reference runs by:
•
Entering a run number series (380452) OR
• In a graph (one often uses the
diagnostics graph), right – clicking a
marquee and choosing ‘Set Airbeam
Reference’ from the options.
It is best to choose runs near a calibration period.
If you are analyzing data with v/w mode switching, select reference runs
with both v and w mode runs in it.
Complete Analysis > Airbeam correction
Go to the PToF tab and make sure the Dva and DC
markers checkboxes are unchecked.
Press the gold Do Corrections button.
A graph of the airbeam
correction factor will appear.
The airbeam correction factor should be near 1.
It’s use in subsequent analyses is chosen by checkboxes in other tabs.
Complete Analysis > Airbeam correction
Airbeam Results:
root:diagnostics:corr_fact
Before pressing leaving the airbeam tab section, make
sure that you have a good airbeam correction factor.
Complete Analysis > PToF Corrections
In the Corrections tab, press the PToF subtab.
There are 2 sections:
DVa
DC marker
Performing the DVa or DC marker corrections can happen at any
time after pre-process.
PToF DVa Discussion
How do we convert PToF bin # to time, then size?
One run of PToF stick data set is a matrix.
Let columns represent m/z dimension (0 – 500 amu),
rows the different PToF size bins (90).
integer amu
Mass 4
1
2
3
90
1
2
3
4
5
6
500
0.0
0.0
0.1
.
.
4.0
1.1
1.0
0.1
0.2
0.2
1.2
0.5
0.7
2.5
0.4
0.3
2.1
0.3 …
0.6 …
3.3 …
0.4
0.1
0.3
0.1
0.5
0.0
0.0
0.1
0.0
PToF stick, Hz
bin number
0
20
100
40
60
80
10
1
3
0.1
0.01
0.001
100
200
300
m/z, amu
400
500
4
5
6
7
PToF DVa Discussion
The formula for converting PToF time to size is:
DVa = dx*((((vg - vl) / (vp - vl)) - 1) ^ (1/b))
where vp=lc/t,
dx, vg, vl, and b are inputs from the daq.
Eqn A
The formula for converting PToF bin number to time is:
PToF time = PToF_t0 + (bin number) * (PToF_step) Eqn B
PToF_t0 incorporates any initial delays
(delays from when chopper is open, first bin measurement, chopper slit center)
PToF_step is a PToF bin width in units of time
(consider coAdds)
If doing calculations by hand, one needs to pay attention to units.
PToF DVa Discussion
In practice, how does this work?
Measurement
time (sec)
integer amu
bin number
1
1
2
3
2
3
0.0 1.1 0.2 …
0.0 1.0 0.2 …
0.1 0.1 1.2 …
500
0.4
0.1
0.3
0.000327
0.000487
0.000647
Eqn B
90
4.0 0.1 0.5 …
PToF Size
(nm)
0.0
1.0465
2.2553
4.4098
Eqn A
0.014567
We don’t change the PToF stick matrix!
1390.4423
PToF DVa
One can set values in this window (blanks
mean keep daq values) OR
Press the simple graph aid button and set
values here (one can explicitly see the
daq values and the DVa calculation).
PToF DVa
PToF DVa Results:
4 parameters vl, vg, dx, b
Often these do not need to be changed from the
settings as recorded by the daq.
Before leaving the PToF DVa section make sure that
these values are to your liking.
The values will not be changed until the gold Do
Corrections button is pushed!
PToF DC Markers Discussion
One run of PToF stick data set is a matrix.
Let columns represent m/z dimension (0 – 500 amu),
rows the different PToF size bins (90).
integer amu
Mass 4
1
2
3
90
1
2
3
4
5
6
500
0.0
0.0
0.1
.
.
4.0
1.1
1.0
0.1
0.2
0.2
1.2
0.5
0.7
2.5
0.4
0.3
2.1
0.3 …
0.6 …
3.3 …
0.4
0.1
0.3
0.1
0.5
0.0
0.0
0.1
0.0
PToF stick, Hz
bin number
0
20
40
60
100
10
80
1
0.1
3
0.01
0.001
100
200
300
m/z, amu
400
500
4
5
6
7
PToF DC Markers Discussion
Mass 40
Define 2 regions.
0
Region 1 Find the average of the ptof sticks (for
avg1
that mass) at each region.
Find the average of both regions, and
subtract this value from the entire
mass size distribution trace.
40
0
60
Avg = (avg1 + avg2)/2
Mass Size distribution -= Avg
Region 2
80
avg2
0.4
20
bin number
bin number
20
40
60
0.8
PToF stick, Hz
(size distribution at a mass)
80
0.0 0.4 0.8
PToF stick, Hz
PToF DC Markers Discussion
In practice how does this work?
Mass 28
0.0
0.0
0.1
4.0
1.1
1.0
0.1
0.1
0.2
0.2
1.2
0.5
0.5
0.7
2.5
0.0
0
0.4
0.6
3.3
0.1
bin number
bin number
integer amu
Mass 67
20
0
40
20
60
40
80
60
40
50
80
PToF stick, Hz
3
4
5
6
7
For each run, each mass (column), find 2 averages,
subtract a value from this column
This is very time consuming!!
Complete Analysis > PToF DC Markers
One can set values in this window OR
Press the simple graph aid button and set
values here.
One can set the start
and stop values for
region 1 and 2 in PToF
bin or time units.
The same regions will
be used for all runs
and all masses except
those indicated.
PToF DC Markers
Results:
Altered PToF stick matrix
Before leaving the PToF DC marker section make sure
that these values are to your liking.
The values will not be changed until the gold Do
Corrections button is pushed!
Frag Checks Discussion
The loadings for most species are the sum of the fractional
mass (fragment or frag, for short) of several integer sticks.
Example: K has loading from the mass at 39 and at K’s isotope at 41 amu. It’s
mass loading is a sum of the fractional mass at 39 and 41.
Mass loading of K = 1*(entire peak at mass 39) + 0.0722*(entire peak at mass 39)
Press ‘Review Frags’ button.
This table encapsulates the linear combination of masses used to derive loadings.
Formulas for some species (such as K) are unambiguous. Other formulas (such
as nitrate) are complicated. Those with experience (!) can tweak the frag table to
reflect conditions which are more consistent with their data.
Complete Analysis > Frag Checks
Press the Checks tab.
Review the species in the list
and the airbeam checkbox.
Press gold ‘Calculate…’ button.
One time-series wave for each species will be
created. They are calculated in the same way as
those found via the MS-tab, time series button.
Frag Checks
Leave the Color by and
Diagnostic menus as
default
Press ‘Plot…’ button.
Review results.
As one become more familiar
with fragmentation patterns,
one can view these graphs to
test the consistency with entries
in the frag table.
Squirrel Analysis Flow Charts
Complete Analysis
HDF Index Tab
Get Index
Pre-Process
Check m/z calibration
m/z fitting params
Check baseline
baseline fitting params
Modify SI
Ion_user
Pre-Process
The corrections and frag
checks analysis steps are
often iterative with the time
series and other graphs .
Airbeam
size dist,
mass spec,
image, etc.
corr_fact
Do Corrections
PToF
time series
avgs, etc.
Frag Checks Tab
size dist,
mass spec,
image, etc.
Corrections Tab
_p data (recalc sticks)
MS Tab
time series
avgs, etc.
PToF Tab
MS Tab
_p data (DAQ sticks)
Get Index
PToF Tab
HDF Index Tab
Quick Look
_p data (dc marker)
Calc Loadings
Loading waves
Tweak
Frag table
Diagnostics
Plot