Data-dependent MSn scan - Data-dependent MSn scan - Orbitrap Tribrid Apex Small Molecule Method Editor Help - Orbitrap Apex Small Molecule MS - Orbitrap Tribrid Apex Small Molecule Method Editor Help - Data-dependent MSn scan - Data-dependent scans - topic

Isolation Mode

Select from the following isolation modes:

• Quadrupole (default): Quadrupole isolation is faster and is done in parallel with analysis, giving the highest instrument duty cycle.
• Ion Trap: Ion trap isolation is fundamental to all MSⁿ analyses (n > 2). Higher-order MSⁿ experiments utilize the ion trap to isolate precursor ions prior to subsequent fragmentation steps.

Isolation Window (m/z)

Specify the isolation window for the precursor ions. The allowable isolation window widths differ for ion trap isolation and quadrupole isolation.

• Range: 0.4–2,000; default: 0.4 (Quadrupole)
• Range: 0.2–2,000; default: 2 (Ion trap)

The mass range for the peak is centered at the precursor mass and ranges from one-half of the isolation window to either side of the precursor mass.

Narrower windows may result in higher selectivity with fewer interfering ions. Wider windows may result in higher sensitivity as more ions enter the MS per unit time.

NOTE: A value that is too low may result in a loss of sensitivity because not all of the ions in a mass peak or in neighboring isotopic peaks are isolated. A value that is too high may cause interference from neighboring peaks.

Isolation Offset

Specify the isolation offset from the following options:

• Off: The isolation window is centered at the most abundant peak of the isotope cluster.
• Isolation is performed at the original precursor mass plus an offset.
• The original isolation window was:
  { precursor – width/2, precursor + width/2 }
• The isolation window with the offset is:
  { precursor + offset – width/2, precursor + offset + width/2 }
• m/z Offset: The mass range for the peak is centered at the precursor mass plus offset and ranges from one-half of the isolation window to either side of the precursor mass plus offset.
• Da Offset: The m/z is calculated using the defined Da offset and the charge state of the detected precursor.
• Isotopic Variant Derived Offset: This option is only available when a ddMSⁿ scan is below an RTLS filter. When selected, the isolation is shifted according to the value of the m/z offset property associated with the compound identified by Real-Time Library Search (“massOffset” property in the spectral library).

Isolation Offset (m/z)

(Available when the Isolation Offset is set to "m/z Offset".)

Specify the offset value (in m/z units) that is applied to the isolation and activation windows relative to that determined through the data-dependent criteria.

For example, if the instrument selects a peak at m/z 500 and the user specifies an m/z 2 width with an m/z 0.5 offset, the isolation range is m/z 499.5–501.5 instead of m/z 499–501 (without the offset).

Range: -200 to 200; default: 2

Isolation Offset (Da)

(Available when the Isolation Offset is set to Da Offset.)

Specify the offset value (in Da units) that is applied to the isolation and activation windows relative to that determined through the data-dependent criteria.

For example, if the instrument selects a peak at Da 500 and the user specifies a Da 2 width with an Da 0.5 offset, the isolation range is Da 499.5–501.5 instead of Da 499–501 (without the offset).

Range: -200 to 200; default: 2

Reported Mass

(Available when the Isolation Offset is set to either "m/z Offset" or "Da Offset".)

Set the reported mass to one of the following options to define which mass is written to the raw file.

• Offset Mass: Offset mass is the center mass of the executed isolation.
• Original Mass: Original mass is the mass that was detected. For example, the mass that would be written to the raw file if the Isolation Offset were set to Off.

For example, if the instrument selects a peak at m/z 500 and the user specifies an m/z 2 width with an m/z 0.5 offset, the isolation range is m/z 499.5–501.5 instead of m/z 499–501 (without the offset).

In this example, the original mass is 500 and the offset mass is 500.5.

Activation Type

Select how the precursor ion is activated for fragmentation.

For more information, see Activation parameters.

Detector Type

Specify the detector type from the following options:

• Ion Trap: Highest sensitivity
• Orbitrap: Highest resolution and mass accuracy.

Ion Trap Scan Rate

(Available when the Ion Trap detector type is selected.)

Faster ion trap scan rates have lower resolution. Specify the ion trap scan rate from the following list:

• Turbo: Uses the linear ion trap accelerated scan mode (125,000 Da/sec). When you select turbo scan, the ion trap performs more scans per unit time at lower resolution. This operating mode is useful for increasing the number of scans across a chromatogram peak during a full-scan type, MS scan mode experiment. This increases the total ion count and increases the signal-to-noise ratio for experiments when instrument noise is present.
• Rapid (default): Uses a scan rate (66,666 Da/sec). which is faster than the normal scan. This scan rate is recommended for fast chromatography.
• Normal: Uses the normal scan rate (33,333 Da/sec) and resolution settings.
• Enhanced: Uses the enhanced scan rate (10,000 Da/sec) and resolution settings. This scan rate is an intermediate scan rate between normal and zoom scan. It provides charge state identifications up to charge state 3, which are typically observed with proteomics samples. However, the scan rate is higher than zoom scan, so the scanning throughput is higher than running zoom scan to accomplish this task.
• Zoom: Uses the zoom scan rate (2,222 Da/sec) and resolution settings. Zoom provides information about the charge state of one or more ions of interest. Zoom scans are slower scans with higher resolution than normal scans.

Scan Range Mode

Select the scan range mode to determine the scan range.

• Auto: The last mass is automatically set to the precursor × charge + 10.0. The first mass is set to the lowest value that retains ions at the last mass.
• Define m/z range: Explicitly define the first and last mass.
• Define First Mass: Explicitly define the first mass, and automatically set the last mass to precursor × charge + 10.0.

Scan Range (m/z)

(Available when the Scan Range Mode is set to "Define m/z range".)

Specify the m/z range over which the ion trap or Orbitrap mass analyzer detects ions. The allowed scan range (in m/z units) depends on the selected mass range and includes the following option(s):

Normal range (Orbitrap and Ion trap): 40–2,000; default: 150–2,000
High range (Orbitrap): 100–6,000; default: 150–2,000
High range (Ion trap): 100–4,000; default: 150–2,000

TIP: As the system optimizes transfer of ions for the defined mass range, it is recommended to restrict the scan range to the region of interest, using a first mass 1 m/z lower than the lightest ion of interest. In addition to the application mode-based defaults, the system templates have recommended application-specific scan ranges.

First Mass (m/z)

(Available when the Scan Range Mode is set to Define First Mass.)

Specify the first mass (lowest m/z) that will be recorded in the spectra.

Normal Range: 40–2,000; default: 100
High Range: 100–6,000; default: 100

AGC Target

The automatic gain control™ (AGC) target value controls the number of ions that are injected into the Orbitrap or Ion trap mass analyzer for mass analysis.

Select from the following options:

• Standard: When Standard is selected, the system sets the recommended target in an automated fashion per scan type and user-defined settings.
• Custom: When Custom is selected, the user defines the target. This selection enables the Normalized AGC Target (%) option.

Absolute AGC Value

The Absolute AGC Value is a conversion of the AGC percentage target set in the method into absolute values. It is a read-only value and is automatically updated according to the scan type and the target percentage.

Normalized AGC Target (%)

(Available when the AGC Target is set to Custom.)

Specify the automatic gain control (AGC) target. This is a percentage representing the maximum number of charges to accumulate for a given analysis.

Range: 0.001–10,000; default: 1,000

The normalized AGC target value is represented as a percentage to aid in calculating the desired AGC target. The base normalized value is different for MSⁿ Level and Detector Type. The values are as follows:

• Scan Type: Full Scan
• Normalized Base (100%), Orbitrap detector = 4e5
• Normalized Base (100%), Ion Trap detector = 3e4
• Scan Type: SIM Scan, MSⁿ Scan
• Normalized Base (100%), Orbitrap detector = 5e4
• Normalized Base (100%), Ion Trap detector = 1e4

The ion routing multipole (IRM) fills ions until it reaches the AGC target or the maximum injection time. The MS then transfers the ions to the Ion Trap or Orbitrap for analysis.

Some scan types allow multiplexing.

• For quadrupole isolation multiplexing in tSIM, the total AGC target is per ion species of a multiplexed group. Each species will inject for different times in order to achieve target.
• For quadrupole isolation multiplexing in tMS² or for ion trap isolation multiplexing, each precursor receives the same injection time.

NOTE: The dynamic range of tMS² multiplexing is more limited than tSIM multiplexing because the injection times of all precursors are equal. Also, Ion Trap isolation multiplexing is in general much less well developed and should be regarded as an experimental feature.

When "Standard" is selected for AGC target, the system will set a recommended value for the scan type. This may deviate significantly from the default 100% Normalized level.

Maximum Injection Time Mode

Select the mode for the maximum injection time from the following options:

• Auto: When Auto is selected, the system calculates the maximum amount of injection time available to maximize sensitivity while maintaining the maximum scan rate.
• Dynamic: When Dynamic is selected, the system calculates the maximum amount of injection time available to maximize sensitivity while maintaining the desired cycle time.
• Custom: When Custom is selected, the user defines the maximum amount of injection time. This selection enables the Maximum Injection Time (ms) option.

Maximum Injection Time (ms)

(Available when the Maximum Injection Time Mode is set to Custom.)

Specify the maximum injection time (in milliseconds) allowed to reach the AGC target.

Range: 0.001–8,000

The ion routing multipole (IRM) fills ions until it reaches the AGC target or the maximum injection time. The MS then transfers the ions to the Ion Trap or Orbitrap for analysis.

Some scan types allow multiplexing.

• For quadrupole isolation multiplexing in tSIM, the total AGC target is per ion species of a multiplexed group. Each species will inject for different times in order to achieve target.
• For quadrupole isolation multiplexing in tMS² or for ion trap isolation multiplexing, each precursor receives the same injection time.

NOTE: The dynamic range of tMS² multiplexing is more limited than tSIM multiplexing because the injection times of all precursors are equal. Also, Ion Trap isolation multiplexing is in general much less well developed and should be regarded as an experimental feature.

Microscans

Specify the number of microscans per scan.

A microscan is one ion injection followed by ion detection. The MS sums microscans to produce one scan, which improves the signal-to-noise ratio of the mass spectral data.

Range: 1–5,000; default: 1

NOTE: The overall scan time increases linearly with the number of microscans, significantly slowing the rate of spectral acquisition.

Data Type

Specify how to collect data during the currently selected scan event using the following options:

• Profile: The profile data type is represented by a collection of signals over several scans. The advantage of profile data is it is easier to classify a signal as a true peak from noise off the instrument.
• Centroid (default): The centroid data type replaces detected peaks with a single m/z and intensity pair, which results in smaller file sizes.

Scan Description

Type an optional description for the selected scan node. This scan description then appears in the scan header of the spectrum’s raw data file.

You can leave the field blank. However, if used, the scan description must be unique within the method and have a maximum of 16 characters. You can use alphanumeric characters, commas, hyphens, parentheses, periods, plus signs, spaces, and underscores.