Below: Elk fibrils imaged at liquid nitrogen temperature (left) in standard bright field TEM imaging and (right) with hole-free phase plate. Sample courtesy of H Young and JP Glaves (biochemistry, University of Alberta). Data was taken by Julie Qian, NINT Electron Microscopy.Microscope phase plates were first developed and demonstrated for light microscopes around 1940 by Dutch scientist, Frits Zernike, rewarded with a Nobel Prize in Physics in 1953 for his work, reports LabCanada. But phase plates are still evolving. In transmission electron microscopy (TEM), the plates improve contrast transfer and lower doses needed to acquire images,  important for imaging organic materials.

TEM phase plate development was extensively pursued by Prof Nagayama's lab in Japan for over a decade. Prof Chiu, Baylor College of Medicine successfully applied the phase plate system on his Omega filtered TEM (JEM-2200FS) to the molecular structure characterisation for proteins.

TEM phase plates however are difficult to produce, as the fabrication process is complicated and delicate. Today, the most successful are made of a uniform and contamination-free carbon thin film of precise thickness with a one micron hole drilled into it.

The biggest difficulty is the need to prevent charging of the carbon phase plate film when in use. Alternatively, a micro-machined electrostatic lens is being tested, potentially offering an additional degree of freedom of the contrast enhancement by controlling the amount of phase shift.

L2R Marek Malac, Masa Kawasaki, Julie Qian, Hiromitsu Furukawa, Miyoko Shimizu, Brian Legg.

A new approach currently offers a potential solution. Hole-free phase plates have been co-developed and patented by a team composed of Dr Marek Malac, a principal investigator at Canada's National Institute for Nanotechnology (NINT), JEOL's Dr Masa Kawasaki, TEM applications scientist, Prof Ray Egerton from NINT and Physics University of Alberta and Dr Marco Beleggia from Denmark Technical University while visiting NINT.

NINT is a government lab, a partnership between Canada's National Research Council, the University and the Government of Alberta. The research team is investigating hole-free phase plates with the JEOL JEM-2200FS Field Emission TEM with an Omega filter, having discovered that Zernike-like phase contrast imaging is easily achieved using a uniform thin film placed in the back focal plane. Using their design, the phase plates are remarkably simple to fabricate and operate. 

Installed in the TEM column, the incident electron beam locally charges the carbon film, leading to phase plate-like effect. Since the electron beam itself is used to charge the carbon film, the centre of the phase plate is determined by the nearly arbitrary position of the electron beam, eliminating the need for precise alignment.

Alternatively, the strong primary electron beam can be used to fabricate the hole in situ eliminating the expense of pre-fabrication, and replacing the need for precise alignment by a much less stringent requirement of low drift of the phase plate mechanism.

"We have been involved with phase plates for just about two years so we are pretty much newcomers. There are groups that have been doing this for a long time and doing very well, but we got success by simplifying the technology and making it somewhat easier to set up and use. Simplification of operation is a major driver for widespread use of the phase plate," said Dr Malac, who heads a team of about 10 members, and continues to work with his advisor, (left) Prof Ray Egerton, expert on electron energy loss spectroscopy (EELS).

The simplified phase plate hardware can easily be inserted to optimise the electron optics of the TEM, according to Dr Kawasaki.

"It's very easy to operate compared to other designs where you have to put the beam onto a centre of a 1 micron hole," he said.
"This phase plate has no hole so the beam can be anywhere around the centre of a phase plate. There are two places in the column where phase plates can be positioned. One is the objective lens area with the specimen just above.

"The other area is the selected area aperture (SAA) position which allows more freedom. At NINT, the hole-free phase plates are installed at this position which is much easier to implement and may avoid a contamination issue that may occur under the environment near the specimen."

"You just put the phase plate in the back focal plane and wait for charge to settle down, then take the data," Dr Malac added. 

Now to investigate how to control the charge amount and the resulting phase shift.