Diagnostic Method Employs Birefringence of Liquid Crystals

A detection system that uses birefringence as the sole optical output signal holds promise as a low-cost, rapid diagnostic tool for identifying biomarkers, viruses, bacteria and parasites in the field.

Scientists from ETH Zurich (the Swiss Federal Institute of Technology) used the phenomenon of birefringence of polarized light from lipid-based lyotropic liquid crystals, which consist of self-assembled structures of fat molecules in water.

A birefringence pattern of a sample positive to Ebola infection. Courtesy of Jijo Vallooran/ETH Zurich.

The diagnostic setup is relatively simple: A drop of blood is placed on a carrier substance. After a few minutes, the slide is placed on a device that emits polarized light using an inexpensive polarization filter. It is covered with a lid containing a second polarization filter, which blocks the light from all materials except crystals or materials with directional properties.

If light is visible through the cross-polarizer filter, a positive diagnosis is made. It is also possible to measure the light intensity, and thus the amount of a given pathogen, through a simple light meter connected to a smartphone.

Lyotropic liquid crystals organize themselves into networks with unique symmetry, which means that their basic motif repeats itself periodically. In the case of liquid crystal cubic phases, the channels are made of lipid bilayer membranes in water and have a diameter of just a few nanometers, so only a few free water molecules are available in the liquid crystal, whereas the majority are bound to the channel walls.

These liquid crystal cubic phases are isotropic, meaning they do not have any birefringent properties. As such, if a slide with a layer of lyotropic liquid crystal films is placed under a light source that allows polarized light to pass through, it appears black when observed through another polarizer tilted at 90°.

To achieve birefringence and thus receive a signal, the researchers added certain enzymes to the liquid crystal to allow a chemical reaction to take place in the nanotubes. Since only a small amount of water is freely available in the nanotubes, the products of the reactions precipitated together to form crystals, which are themselves birefringent.

A closer look at the sample through a second polarization filter placed above it and perpendicular to the first showed a light pattern in instances where the enzyme had reacted with the substance tested.

"This birefringence pattern is the only signal that we need to use for diagnostics and analysis," said professor Raffaele Mezzenga.

At the beginning of their research, the scientists tested their system with chemical compounds that could be enzymatically converted. They then adapted their method for medically relevant substances, such as glucose and cholesterol. In further steps, they broadened the method to test for bacteria and viruses, starting with HIV. Eventually the team was able to show that the method could be adapted to diagnose malaria caused by Plasmodium parasites.

"Plasmodium parasite invades erythrocytes and digests hemoglobin," said Mezzenga. "The heme component, which is toxic to the parasites, is crystallized and thus has inherently birefringent surfaces. So it's not necessary to mark it with antibodies and no enzymatic reaction is required."

Normally, viruses and bacteria must be made visible and chemically active with specific antibodies with enzymes coupled to them before they can be detected by birefringence.

"Our test system can be extended to a large number of different viruses or bacteria. It is totally flexible," said researcher Jijo Vallooran. "Other than a refrigerator to store the antibodies and enzymes, the user needs only the box to detect the polarized light and the lipid carrier substance. This is very inexpensive."

Pathogens such as HIV or Ebola can be detected very rapidly, and a reliable result received within less than an hour, the researchers said.

"Our technology is very suitable for use in the field and the early detection of diseases," said Vallooran.

The findings were published in Advanced Functional Materials (doi: 10.1002/adfm.201503428).