MIMS Acquisition Protocol

As described in Section 2.3.2, the MIMS system has two primary components: the laser ablation (LA) system on the front end and the spectrometer on the back end. For the applications in this work, the back end spectrometer was either an optical emission spectrometer (OES) or an inductively coupled plasma mass spectrometer (ICP-MS). Without loss of generality, the back end spectrometer instrument will be referred to as a mass spectrometer (MS) in this Chapter. To acquire a dataset, the first step is to provide the instructions in the form of an∗.lzs file to the LA system. To

produce these instructions, CETAC’s DigiLaz III software is used in conjunction with a custom MATLAB graphical user interface (GUI), developed by Boston University undergraduate students Daniel Brewster and Casey Kurosawa. The GUI facilitates the construction of more complex data acquisition routines. Fundamentally, the Dig- iLaz III software provides a GUI through which the user can interact with live video footage to construct an imaging routine. Figure 4·1 shows a screen shot of the Digi- Laz III software GUI. The information inputted using DigiLaz III populates an excel table which DigiLaz III uses to generate an ∗.lzs file. The parameters defined using

61 Spatial Specifications (columns 1 - 11)

Scan Name X1 X2 Y1 Y2 Z1 Z2 Method Aperture Size Space Between

Spots Space Between Lines string [µm] [µm] [µm] [µm] [µm] [µm] category [µm] [µm] [µm] Nominal parameter values – – – – – – Single Line Scan 20 0 20

Laser Specifications (columns 12 - 17)

Energy Pulse Rep Rate Scan Rate Number of Shots Defocus Defocus Amount

[%] [Hz] [µm/s] integer Y/N [%]

5 20 60 N 0

Gas and Timing Specifications (columns 18 - 25) He Flow Rate Pause Between Samples Shutter Delay Gas Blank Trigger Delay Sample Run Time Total Sample Time Number of Runs

[L/min] [sec] [sec] [sec] [sec] [sec] integer

The optical system that is part of the CETAC LA instrument has limited field of view; therefore, to generate an LA routine, the DigiLaz III software first constructs an image of the ablation cell by stitching a 12 × 8 grid of smaller field of view images together (see Fig. 4·1, bottom right). This mosaic image is representative of the entire 39.6 × 39.6 mm2 _{ablation area within the cell. The image can be saved and}

used to identify arbitrary stage coordinates within the LA cell. Our group’s custom MATLAB GUI takes this mosaic image as an input and allows the user to indicate multiple regions of interest within the ablation cell (see Fig. 4·2). Using this map between pixels in the mosaic image and stage positions, the MATLAB GUI allows the user to draw regions of interest on the mosaic image to generate an ∗.xls file

with the appropriate parameters. The∗.xls file can be uploaded into the DigiLaz III

software to generate a correctly formatted LA protocol. The MATLAB GUI facilitates the synthesis of an LA routine in three respects: 1) it enables guided and automated naming of the lines, 2) it allows the user to quickly indicate imaging regions that may be spatially distant with respect to the field of view of the live video, thus eliminating the need to reposition the stage in order to get an appropriate field of view, and 3) it is easier to identify distinct samples within the cell since the full imaging region is the focus rather than the live video field of view being the focus. The MATLAB GUI also allows for specific imaging sequences, such as calibration or standard regions, to be repeated automatically.

In addition to parameterization of the LA instrument, operation parameters for the MS instrument must also be specified. Each instrument is accompanied by a manufacturer-provided software which enables the specification of acquisition pa- rameters including isotope selection, instrument sensitivity, isotope sampling time parameters, among others. These parameters can be adjusted to fit experimental requirements and constraints. An example of the an MS GUI is depicted in Fig. 4·3.

Figure 4·2: MATLAB graphical user interface for constructing laser ablation routines.

Figure 4·3: Example of manufacturer’s graphical user interface for the specification of parameters of the Element ICP-MS instrument.

Figure 4·4: Timing diagram showing relative timing of the laser (top), the spectrometer (middle), and the amount of particles generated from the laser ablation cell (bottom) for two lines of data. [Blue area] MS begins sampling data once trigger signal is received from the LA instru- ment (blank period). [Red area] Laser fires on the sample and generates particles in the ablation cell. [Green area] Laser turns off, but the MS continues to sample data (washout period). [Purple area] Time delay between lines inserted to allow both the LA instrument and the MS to prepare for the acquisition of a new line of data (no data).

The LA and MS communicate via an electrical trigger signal. The trigger signal indicates the start of a line of data in the metallomic image and prompts the MS to begin acquiring data for the indicated isotopes. Each line begins with a laser off period where the shutter is maintained on the laser, but the MS begins sampling the detector. A laser on period starts during which particles are generated from the sample. Once the laser on period ends, the spectrometer continues sampling for the

total user defined period. A time delay between lines, where neither the laser nor the spectrometer are active, is incorporated to ensure proper synchronization of the instruments. The timing of the trigger and data acquisition events are depicted in Fig. 4·4. For the MS instruments, the data for all indicated isotopes are saved in a text file with an instrument-specific formatting. Data acquisition was primarily conducted by analytical chemists Noel Casey, Ph.D., and Bo Yan, Ph.D.