Underwater Atomic Force Microscopy

To fully understand the structure and electrical properties of biological materials such as cell membranes or proteins, biologists need to study them in their native, watery environments. Now researchers report a technique that could provide this information at high resolution under biologically friendly conditions. The team has developed a kind of atomic force microscopy that works on samples sitting in water and that is gentle enough to analyze fragile biological surfaces (Langmuir 2013, DOI: 10.1021/la4002797).

In a typical AFM experiment, a sharp tip scans over a surface, producing an image based on the forces the tip experiences as it interacts with molecules or atoms on the surface. AFM can generate atomic-scale information on the topography and electrical properties of a surface. The technique works well in air and with robust samples that can withstand contact with the hard, sharp imaging tip.

Biological structures like cells and proteins are wet and squishy—not a natural fit for AFM. Seong H. Kim, a chemical engineer at Pennsylvania State University, wants to make an AFM technique that is compatible with biological samples. He thinks that such a technique could help biologists study proteins embedded in cell membranes or the electrical potentials of nerve cells.

To adapt AFM to biological samples, Seong built on a method called scanning polarization force microscopy that was developed in the 1990s by researchers at the Lawrence Berkeley National Laboratory (Appl. Phys. Lett. 1995, DOI: 10.1063/1.114541). In that method, an AFM tip under an applied voltage maps electrical charges on a surface exposed to air. The method does not require physical contact with the surface being imaged. Instead, static charges on the surface either attract or repel the tip, creating a measurable force.

Researchers in the field thought the technique wouldn’t work underwater. And when Kim’s group first tried the method in water, they found that dissolved ions coated the AFM tip, interfering with how the tip interacted with a sample’s surface. But they overcame this problem by oscillating between positive and negative voltages at the tip. With these oscillations, the ions couldn’t build up fast enough to interfere with the measurements.

To prove the concept of the underwater method, Kim’s group imaged a gold surface covered with self-assembled monolayers of charged polymers. With the AFM technique, the team could make a map of the surface’s topography and distinguish between positive and negative charges.

Kim says it’s not yet clear what the ultimate resolution of this wet method will be. The best AFM methods can provide atomic resolution. Right now Kim says his method can discern objects as small as 200 nm. The imaging tips the team used are about 10 to 20 nm wide, so greater resolution should be possible.

“The fact that you can truly operate in a liquid with this method could make it interesting for biological researchers,” says Adam Z. Stieg of the California NanoSystems Institute at the University of California, Los Angeles. If Kim’s group can demonstrate it with actual biological samples and can make a user-friendly version, he says, the method could offer something unique for biologists. No technique currently used could match the spatial resolution possible with AFM, Stieg says.