ITMO Scientists' Collaboration with German and Swiss Colleagues Wins ERA.Net Research Grant

A team of scientists from ITMO’s International Research and Education Center for Physics of Nanostructures, together with their colleagues from the Federal Institute for Materials Research and Testing (Germany) and ETH Zurich (Switzerland), have been awarded a grant by the European initiative ERA.Net RUS Plus. Their three-year project concerns the modeling and design of new functional nanomaterials with potential applications in biomedicine and diagnostics. As the researchers note, nanomaterials based on ternary quantum dots can be used to create a sensor platform with unprecedented filtering and sensitivity capabilities for the purpose of multiplexed single-cell pathogen analysis. In the future, results of this research can contribute to the development of more precise and compact devices used for fluorescence analysis of nanostructures.

What is ERA.Net RUS Plus

ERA.Net RUS Plus is an international initiative aimed at establishing long-term scientific and technological partnerships between member countries of the European Union, EU-associated countries and Russia. One of the project’s key objectives is the inclusion of Russian scientists in the European Research Area (ERA).

Unlike the European framework program Horizon 2020 (learn more about it here), ERA.Net RUS Plus offers a closer collaboration with Russian scientists. The initiative was launched in 2014; every three years, a competition is held to select the best projects from the program’s participant countries.

Applications are accepted if they involve fundamental research projects by scientists or teams from Russia, Belgium, Bulgaria, Germany, Latvia, Moldova, Romania, Serbia, Slovakia, Turkey, Finland, Switzerland and Estonia. Among the common topics are the five areas of research vital to the scientific and technological development of countries-participants of ERA.Net RUS Plus:

  1. Nanotechnologies (advanced nanosensors for environmental and health monitoring; new nanomaterials);
  2. Environment protection and climate change (effect of climate change and extreme weather on the environment, prevention and clean-up of water ecosystem contaminations);
  3. Health science (restorative medicine, biomaterials and organ-on-a-chip systems, treatments for oncological, cardiovascular and infectious diseases);
  4. Humanities and social sciences (demography, conflicts and security issues, traditional and non-traditional cultural values; goals and opportunities for regional development and social harmony);
  5. Robotics (human-machine interaction, agricultural, medical, industrial, nautical and educational robots).

Each project is submitted to the competition by a consortium that includes scientific groups from at least three countries; each group receives support from their own country. Russian scientists receive funding from the Ministry of Education and Science, the Russian Foundation for Basic Research, and the Ural and Far East branches of the Russian Academy of Sciences.

The completion time for projects submitted to the competition is 2 to 3 years. The competition’s expert council and independent experts evaluate the applications.

In 2017, more than 200 projects applied to the program; 28 were chosen by ERA.Net’s group of sponsoring organizations to receive funding to a total of 11 million euros. One of the winners in the nanotechnologies area was the project “Modeling-aided design of a ternary quantum dot-based platform for multiplexed cell analysis”, developed by a consortium including teams from ITMO University, the Federal Institute for Materials Research and Testing (Germany) and ETH Zurich (Switzerland), which is among the world’s top 10 universities according to Times Higher Education (THE) rankings.

As Alexander Baranov, professor at ITMO’s Department of Optical Physics and Modern Natural Science and head of the “Optics of Quantum Nanostructures” laboratory, notes, this collaboration with international colleagues is largely a result of work by young scientists who graduated from the department and went on to work at leading universities and research centers abroad.

Alexander Baranov
Alexander Baranov

“Our graduates who work as postdocs at universities abroad have helped us establish collaborations in physics and nanostructure technology with many universities, like Trinity College Dublin, University of Campinas in Brazil and several French universities. This project was initiated by our graduate, Irina Martynenko, who currently works at the Federal Institute for Materials Research and Testing in Berlin. As she discussed promising research areas with her German colleagues, it became clear that we have common interests and we can do joint research in this area. A little later we were joined by scientists from ETH Zurich; there, they have the equipment needed to produce lab samples of fluorescence sensors based on our research,” – says Prof. Baranov.

After being approved by ERA.Net RUS Plus, the project was also given approval by the Russian Ministry of Education and Science, becoming one of the three approved projects from Russia.

Project plans

The joint project is expected to last three years; in that time, the scientific teams plan to model a new sensor platform based on polymeric microparticles doped with non-toxic luminescent quantum dots using AglnS and CulnS semiconductors. The resulting structures can be used to detect pathogen biomarkers in single cells; in the future, research done as part of the project can help create more precise and compact analytical devices – flow cytometers with a system for the detection of spectral and time parameters of biomarkers’ fluorescence.

What is fluorescence analysis of pathogenic microorganisms? This is a well-known method of studying various objects, – starting from human tissue cells and up to foods or geological materials – based on observing their fluorescence. It is used in medical diagnostics (for instance, microsporum-infected tissue can be found by its bright green fluorescence under ultraviolet light) to determine the extent of infection in seeds and plants, to analyze the organic makeup of soil and other purposes.

Fluorescent dyes
Fluorescent dyes

This kind of analysis can consider the objects’ own fluorescence or the fluorescence of special markers applied to it. Different biological structures are marked with fluorescent dyes; then, special analytical equipment (like a fluorescent flow cytometer) measures the number of marked bioobjects in a sample. In its simplest form, it is a capillary tube through which a solution infused with the analyzed objects (a lysate of cells or actual cells) is pumped, while a laser illuminates it and excites the markers’ fluorescence.

The flow cytometers used today are quite sizeable, and one of the team’s engineering challenges is to make them smaller and, if possible, even portable. Moreover, existing versions are not capable of analyzing each individual cell, which means that another task is to increase the devices’ sensitivity, making them more useful when attempting to detect various specific pathogenic microorganisms.

Why is so difficult to ramp up the sensitivity? First of all, the less cells that are to be examined, the less the amount of fluorescence. Secondly, it must be kept in mind that biological structures, just like fluorescent markers, emit light under laser radiation. Therefore, the valid signal of fluorescent markers will be mixed in with the noise from autoluminescence.

Several years ago, a method was suggested as a solution to this issue: to sort signals not only according to their wavelength and intensity, but also its decay time. Autoluminescence usually exhibits a decay time of about 10 nanoseconds – nearly the same as fluorescent markers. The solution suggests to attach so-called donors to each marker; these donors are objects with decay time longer than that of the markers themselves. Through nonradiative energy transfer from the donor to the marker, the latter’s decay time is increased significantly. Using such a method for analysis lets scientists filter out autoluminescence.

In modern scientific practice, glass nanoparticles doped with rare-earth elements are used as donors; these particles’ decay time is measured in microseconds. However, there are several drawbacks to these particles: they are expensive to produce and don’t always match the marker.

“This is the reason why we need to find objects that would exhibit all the properties of a standard donor, i.e. longer luminescence decay time and compatibility with the marker - and yet also be cheap enough,” – explains Prof. Baranov – “In our project, we’ve proposed using quantum dots of particular type for this purpose. We already know how to work with these – they are the so-called trenary quantum dots of the semiconductors AglnS and CulnS. By changing their size and composition, we can adjust their luminescence band to match the absorption band of each marker; at the same time, they are cheap enough. We know how to produce such quantum dots, and produced some in collaboration with our colleagues from Trinity College in Dublin.”

These quantum dots have a few other attractive qualities: for instance, they have a wide luminescence band and the different parts of that band exhibit different fluorescence lifetimes, which means that markers can be sorted both by spectral and time parameters at the same time. Quantum dots of this kind, therefore, are useful in multiplexed cell analysis, where the number of parameters can be increased, making the selection process more precise and selective.

Work on the project will be split into several stages, with tasks distributed among the three teams in Russia, Germany and Switzerland. Researchers at ITMO, for example, will synthesize ternary quantum dots, study their parameters (such as size, shape and elemental composition, as well as spectral and kinetic optical parameters) and model various processes such as that of nonradiative resonant energy transfer.

The project will also involve the creation of polymeric microcapsules with magnetic cores; the capsules’ outer surface will be infused with ternary quantum dots. The use of an external magnet is expected to help increase the concentration of markers in the area being analyzed. Synthesized microcapsules will be sent to the Federal Institute for Materials Research and Testing, where researchers will develop methods of their bioconjugation, and to ETH Zurich – the team there will work on creating a method for detecting luminescence in a flow cytometer.

ETH Zurich
ETH Zurich

It should be noted that Zurich is also where a new method is currently being developed that would make it possible to capture single cells for analysis instead of examining a larger quantity. With that method, the cell is destroyed in order to study its contents, which prevents the common issues of quantum dots penetrating the cellular membrane.

Future prospects

The main goal of the project, notes Prof. Baranov, is the development (theoretical modeling, design and evaluation) of an all-purpose sensor platform base, exhibiting unprecedented filtering and sensitivity capabilities, and based on polymeric microparticles, doped with non-toxic ternary quantum dots. Unlike the currently existing systems, such a platform would make it possible to perform multiplexed single-cell analysis of cell components and pathogens while filtering by the spectrum, intensity and lifetime of their fluorescence, as well as to develop methods of detecting analytes when using miniature flow cytometers.

“One of the main results of this work will be the introduction of this technology here, in Russia,” – adds Alexander Baranov – “We already know how to make quantum dots, we know how to adjust their parameters, how to create various fluorescent markers; moreover, there are already studies expanding on the methods of their bioconjugation, i.e. their interaction with biological target objects. However, there is very little known here about microfluid luminescent flow microcytometers. We think there’s a lot of potential in applying this technology in the form of a unique tabletop device.”

The authors note that this joint interdisciplinary project holds a lot of promise for a multitude of applications.

“Its completion will have a significant effect on the potential for use of fluorescent analysis in two areas – microfluidics and flow cytometry, which are widely used for cell research, biomarker detection in medical diagnostics, and for pathogen analysis,” – say the researchers.

It is also said that the new knowledge acquired as part of this project will later be included in the educational programs of the three universities involved in the project.

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