Nanoscale Characterization of Liquid-Solid Interfaces by Coupling Cryo-Focused Ion Beam Milling with Scanning Electron Microscopy and Spectroscopy

The cryo-SEM and FIB method can be used to study solid-liquid interfaces and biological samples while preserving the samples native structure. The main advantage of this technique is that the cryo-SEM allows the user to quickly probe the interface of macroscopic devices like coin cell battery electrodes with tens of nanometers resolution. Begin by installing a cryo-SEM stage and an anticontaminator.

Evacuate the SEM chamber and adjust the gas injection system, GIS, platinum source so that when inserted, the source will sit approximately five millimeters away from the sample surface. Set the GIS temperature to 28 degrees Celsius and open the shutter to vent the system for 30 seconds to clear out any excess material. Then, allow the SEM chamber to evacuate for a minimum of eight hours.

At the end of the evacuation period, set the microscope and prep stages to minus 175 degrees Celsius and set the anticontaminator to minus 192 degrees Celsius. To vitrify the sample, sequentially fill the main volume of the nitrogen dual-pot slusher and the surrounding volume with liquid nitrogen until the liquid nitrogen stops bubbling. Seal the filled slusher with the lid and initiate the slush pump.

When the liquid nitrogen starts solidifying, begin venting the slush pot. Once the pressure is high enough to allow the pot to be opened, quickly but gently place the sample in the nitrogen. When the boiling has ceased around the sample, use a pre-cooled transfer rod to transfer the sample to the vacuum chamber of a pre-cooled SEM shuttle just before the nitrogen starts to freeze.

Quickly transfer the shuttle to the airlock of the prep chamber and pump on the transfer system. If desired, sputter five to 10 nanometers of a gold-palladium layer onto the sample surface to mitigate charging. Then, transfer the sample shuttle as quickly and smoothly as possible onto the cooled microscope stage.

For sample surface imaging, first image the sample at a 100 X magnification. Next, bring the sample to an approximately eucentric height and acquire a second low-magnification image. Select a sacrificial test region within the vitrified liquid and identify any potential issues that may be present due to beam damage or charging.

Search the sample for the regions-of-interest. When a region has been identified, tilt the sample so that the surface is normal to the direction of the platinum GIS needle and insert the GIS needle. Warm the surface to 28 degrees Celsius and open the valve for approximately 2.5 minutes before retracting the source.

Tilt the sample shuttle toward the focused ion beam source and expose the organo-metallic platinum to a 30-kilovolt ion beam at 2.8 nanoamps and an 800 X magnification for 30 seconds. Then, image the sample surface with the electron beam to verify that the surface is smooth and lacks any signs of charging. To prepare a cross-section, first use the ion beam at 30 kilovolts and a lower bulk milling current of approximately 2.8 nanoamps to acquire a snapshot of the sample surface.

Identify the feature of interest and measure out the rough placement of the cross-section. To create a side window for the x-rays, draw a regular cross section rotated 90 degrees relative to where the trench will be and place the side window with one edge roughly flush with the desired final cross section. Resize the rotated pattern to maximize the number of x-rays to exit the cross section surface.

Use a high current to create a regular cross-section just large enough to reveal the feature of interest and use the ion beam at 30 kilovolts and the current of interest to acquire snapshot of the sample surface. Identify the feature of interest and finalize the placement of the trench. The trench should extend past either side of the feature of interest by a few microns.

Confirm that there is one micrometer of material between the edge of the trench and the desired final cross-section and use the milling application to set the Z-depth to two micrometer, regularly pausing the milling process to image the cross-section with the electron beam as necessary. When the trench is much deeper than the feature of interest, note the amount of time needed to create the rough trench to guide the depth. To create a final, clean cross-section, lower the ion beam current to approximately 0.92 nanoamps and image the sample surface.

After verifying the location of the feature of interest, use the focused ion beam software to draw a cleaning cross-section and overlap the cleaning window with the pre-made trench by at least one micrometer to help mitigate the re-deposition. Then, use the time needed to create the trench to set the Z-depth value. For EDX mapping, select the appropriate beam conditions for the sample and orient the sample to maximize the x-ray counts.

Insert the EDX detector and set the appropriate process time. In the detector software, open the Microscope Setup and start the electron beam image. Click Hit Record to measure the count rate and the dead time.

If the dead time needs to be adjusted, change the EDX time constant. Once the detector conditions have been established, collect the electron beam image and open Image Setup to select the bit depth and image resolution. Select the x-ray map resolution, spectrum range, number of channels, and map dwell time.

The energy range can be as low as the beam energy used. Then, in the EDX software, select the area to map over. When the map is complete, save the map as a data cube.

These images of bare lithium foil milled at 25 and minus 165 degrees Celsius highlight how cooling to cryogenic temperatures can help preserve samples during focused ion beam milling. For EDX experiments, the focused ion beam milling geometry should be optimized and the position of the EDX detector should be taken into account. Here, the difference between a well-and a poorly-prepared cryo-immobilized sample, both using the lithium metal battery as an example, can be observed.

Although both samples were nominally prepared according to the same procedure, a brief exposure to air most likely resulted in the surface reactions observed in the poorly-prepared sample. Mapping a lithium deposit in 1, 3-dioxolane, 1, 2-dimethoxyethane with non-optimal conditions results in contrast variations, likely an indication of an initially well preserved interface that is lost due to radiation damage during mapping. In contrast, this map of dead lithium embedded in vitrified electrolyte and the lithium substrate beneath was performed at two kilovolts and 0.84 nanoamps, preserving the sample surface morphology.

Although some damage is still visible after mapping, the extent of the damage is substantially reduced. In this analysis, EDX mapping was used to locate iron oxide nanoparticles grown in a silica hydrogel. Large field-of-view scans allowed the identification of regions-of-interest, while more localized scans were used for site-specific milling.

Sample charging can be detrimental to the success of this procedure. Remember to lower beam currents and dwell times as necessary to limit the effects of charging. After this, a cryo-FIB lift-out can be performed to prepare a site-specific lamella for TEM analysis.

Samples can be imaged at sub-angstrom resolution and map the chemical distribution using EELS and EDX in a TEM instrument.