Laboratory of Molecular Biophysics

NamePositionContacts
Aleksej Mezin+7(495)336-07-77
Alexander Schapov+7(495)330-61-65
Vladimir Oleinikov, D.Scdepart. dir.voleinik@mail.ru+7(495)335-43-66
Konstantin Mochalov, Ph.D.s. r. f.mochalov@mail.ru+7(495)336-07-77
Yury Kozmins. r. f.zibotic@mail.ru+7(495)330-61-65
Marina Tretyakr. f.marinatretyak@mail.ru+7(495)995-55-57#2113
Daria Solovyevaj. r. f.d.solovieva@mail.ru+7(495)3354366, +7(965)3922491
Tatiana Sukhanovaj. r. f.sukhanovat@mail.ru+7(495)330-61-65
Vasiliy KovalPhD stud.tokojami@ya.ru
1164eng.tosha111@gmail.com
Veronika Manohinaeng.+7(495)330-61-65
Mikhail Birjukoveng.+7(495)330-61-65

Selected publications

  1. Mochalov K.E., Chistyakov A.A., Solovyeva D.O., Mezin A.V., Oleinikov V.A., Vaskan I.S., Molinari M., Agapov I.I., Nabiev I., Efimov A.E. (2017). An instrumental approach to combining confocal microspectroscopy and 3D scanning probe nanotomography. Ultramicroscopy 182, 118–123 [+]

    In the past decade correlative microscopy, which combines the potentials of different types of high-resolution microscopies with a variety of optical microspectroscopy techniques, has been attracting increasing attention in material science and biological research. One of outstanding solutions in this area is the combination of scanning probe microscopy (SPM), which provides data on not only the topography, but also the spatial distribution of a wide range of physical properties (elasticity, conductivity, etc.), with ultramicrotomy, allowing 3D multiparametric examination of materials. The combination of SPM and ultramicrotomy (scanning probe nanotomography) is very appropriate for characterization of soft multicompound nanostructurized materials, such as polymer matrices and microstructures doped with different types of nanoparticles (magnetic nanoparticles, quantum dots, nanotubes, etc.), and biological materials. A serious problem of this technique is a lack of chemical and optical characterization tools, which may be solved by using optical microspectroscopy. Here, we report the development of an instrumental approach to combining confocal microspectroscopy and 3D scanning probe nanotomography in a single apparatus. This approach retains all the advantages of SPM and upright optical microspectroscopy and allows 3D multiparametric characterization using both techniques. As the first test of the system developed, we have performed correlative characterization of the morphology and the magnetic and fluorescent properties of fluorescent magnetic microspheres doped with a fluorescent dye and magnetic nanoparticles. The results of this study can be used to obtain 3D volume images of a specimen for most high-resolution near-field scanning probe microscopies: SNOM, TERS, AFM-IR, etc. This approach will result in development of unique techniques combining the advantages of SPM (nanoscale morphology and a wide range of physical parameters) and high-resolution optical microspectroscopy (nanoscale chemical mapping and optical properties) and allowing simultaneous 3D measurements.

    ID:1985
  2. Efimov A.E., Agapov I.I., Agapova O.I., Oleinikov V.A., Mezin A.V., Molinari M., Nabiev I., Mochalov K.E. (2017). A novel design of a scanning probe microscope integrated with an ultramicrotome for serial block-face nanotomography. Rev Sci Instrum 88 (2), 023701 [+]

    We present a new concept of a combined scanning probe microscope (SPM)/ultramicrotome apparatus. It enables "slice-and-view" scanning probe nanotomography measurements and 3D reconstruction of the bulk sample nanostructure from series of SPM images after consecutive ultrathin sections. The sample is fixed on a flat XYZ scanning piezostage mounted on the ultramicrotome arm. The SPM measuring head with a cantilever tip and a laser-photodiode tip detection system approaches the sample for SPM measurements of the block-face surface immediately after the ultramicrotome sectioning is performed. The SPM head is moved along guides that are also fixed on the ultramicrotome arm. Thereby, relative dysfunctional displacements of the tip, the sample, and the ultramicrotome knife are minimized. The design of the SPM head enables open frontal optical access to the sample block-face adapted for high-resolution optical lenses for correlative SPM/optical microscopy applications. The new system can be used in a wide range of applications for the study of 3D nanostructures of biological objects, biomaterials, polymer nanocomposites, and nanohybrid materials in various SPM and optical microscopy measuring modes.

    ID:1986
  3. Bobrovsky A., Mochalov K., Oleinikov V., Solovyeva D., Shibaev V., Bogdanova Y., Hamplová V., Kašpar M., Bubnov A. (2016). Photoinduced Changes of Surface Topography in Amorphous, Liquid-Crystalline, and Crystalline Films of Bent-Core Azobenzene-Containing Substance. The journal of physical chemistry. B 120 (22), 5073–82 [+]

    Recently, photofluidization and mass-transfer effects have gained substantial interest because of their unique abilities of photocontrolled manipulation with material structure and physicochemical properties. In this work, the surface topographies of amorphous, nematic, and crystalline films of an azobenzene-containing bent-core (banana-shaped) compound were studied using a special experimental setup combining polarizing optical microscopy and atomic force microscopy. Spin-coating or rapid cooling of the samples enabled the formation of glassy amorphous or nematic films of the substance. The effects of UV and visible-light irradiation on the surface roughness of the films were investigated. It was found that UV irradiation leads to the fast isothermal transition of nematic and crystalline phases into the isotropic phase. This effect is associated with E-Z photoisomerization of the compound accompanied by a decrease of the anisometry of the bent-core molecules. Focused polarized visible-light irradiation (457.9 nm) results in mass-transfer phenomena and induces the formation of so-called "craters" in amorphous and crystalline films of the substance. The observed photofluidization and mass-transfer processes allow glass-forming bent-core azobenzene-containing substances to be considered for the creation of promising materials with photocontrollable surface topographies. Such compounds are of principal importance for the solution of a broad range of problems related to the investigation of surface phenomena in colloid and physical chemistry, such as surface modification for chemical and catalytic reactions, predetermined morphology of surfaces and interfaces in soft matter, and chemical and biochemical sensing.

    ID:1987
  4. Ivanov A.A., Koval V.S., Susova O.Y., Salyanov V.I., Oleinikov V.A., Stomakhin A.A., Shalginskikh N.A., Kvasha M.A., Kirsanova O.V., Gromova E.S., Zhuze A.L. (2015). DNA specific fluorescent symmetric dimeric bisbenzimidazoles DBP(n): the synthesis, spectral properties, and biological activity. Bioorg. Med. Chem. Lett. 25 (13), 2634–8 [+]

    A series of new fluorescent symmetric dimeric bisbenzimidazoles DBP(n) bearing bisbenzimidazole fragments joined by oligomethylene linkers with a central 1,4-piperazine residue were synthesized. The complex formation of DBP(n) in the DNA minor groove was demonstrated. The DBP(n) at micromolar concentrations inhibit in vitro eukaryotic DNA topoisomerase I and prokaryotic DNA methyltransferase (MTase) M.SssI. The DBP(n) were soluble well in aqueous solutions and could penetrate cell and nuclear membranes and stain DNA in live cells. The DBP(n) displayed a moderate effect on the reactivation of gene expression.

    ID:1363
  5. Mochalov K.E., Efimov A.E., Bobrovsky A., Agapov I.I., Chistyakov A.A., Oleinikov V., Sukhanova A., Nabiev I. (2013). Combined scanning probe nanotomography and optical microspectroscopy: a correlative technique for 3D characterization of nanomaterials. ACS Nano 7 (10), 8953–62 [+]

    Combination of 3D structural analysis with optical characterization of the same sample area on the nanoscale is a highly demanded approach in nanophotonics, materials science, and quality control of nanomaterial. We have developed a correlative microscopy technique where the 3D structure of the sample is reconstructed on the nanoscale by means of a "slice-and-view" combination of ultramicrotomy and scanning probe microscopy (scanning probe nanotomography, SPNT), and its optical characteristics are analyzed using microspectroscopy. This approach has been used to determine the direct quantitative relationship of the 3D structural characteristics of nanovolumes of materials with their microscopic optical properties. This technique has been applied to 3D structural and optical characterization of a hybrid material consisting of cholesteric liquid crystals doped with fluorescent quantum dots (QDs) that can be used for photochemical patterning and image recording through the changes in the dissymmetry factor of the circular polarization of QD emission. The differences in the polarization images and fluorescent spectra of this hybrid material have proved to be correlated with the arrangement of the areas of homogeneous distribution and heterogeneous clustering of QDs. The reconstruction of the 3D nanostructure of the liquid crystal matrix in the areas of homogeneous QDs distribution has shown that QDs do not perturb the periodic planar texture of the cholesteric liquid crystal matrix, whereas QD clusters do perturb it. The combined microspectroscopy-nanotomography technique will be important for evaluating the effects of nanoparticles on the structural organization of organic and liquid crystal matrices and biomedical materials, as well as quality control of nanotechnology fabrication processes and products.

    ID:1122
  6. Melnikau D., Savateeva D., Lesnyak V., Gaponik N., Fernández Y.N., Vasilevskiy M.I., Costa M.F., Mochalov K.E., Oleinikov V., Rakovich Y.P. (2013). Resonance energy transfer in self-organized organic/inorganic dendrite structures. Nanoscale 5 (19), 9317–23 [+]

    Hybrid materials formed by semiconductor quantum dots and J-aggregates of cyanine dyes provide a unique combination of enhanced absorption in inorganic constituents with large oscillator strength and extremely narrow exciton bands of the organic component. The optical properties of dendrite structures with fractal dimension 1.7-1.8, formed from J-aggregates integrated with CdTe quantum dots (QDs), have been investigated by photoluminescence spectroscopy and fluorescence lifetime imaging microscopy. Our results demonstrate that (i) J-aggregates are coupled to QDs by Förster-type resonant energy transfer and (ii) there are energy fluxes from the periphery to the centre of the structure, where the QD density is higher than in the periphery of the dendrite. Such an anisotropic energy transport can be only observed when dendrites are formed from QDs integrated with J-aggregates. These QD/J-aggregate hybrid systems can have applications in light harvesting systems and optical sensors with extended absorption spectra.

    ID:1128
  7. Generalova A.N., Oleinikov V.A., Sukhanova A., Artemyev M.V., Zubov V.P., Nabiev I. (2013). Quantum dot-containing polymer particles with thermosensitive fluorescence. Biosens Bioelectron 39 (1), 187–93 [+]

    Composite polymer particles consisting of a solid poly(acrolein-co-styrene) core and a poly(N-vinylcaprolactam) (PVCL) polymer shell doped with CdSe/ZnS semiconductor quantum dots (QDs) were fabricated. The temperature response of the composite particles was observed as a decrease in their hydrodynamic diameter upon heating above the lower critical solution temperature of the thermosensitive PVCL polymer. Embedding QDs in the PVCL shell yields particles whose fluorescence is sensitive to temperature changes. This sensitivity was determined by the dependence of the QD fluorescence intensity on the distances between them in the PVCL shell, which reversibly change as a result of the temperature-driven conformational changes in the polymer. The QD-containing thermosensitive particles were assembled with protein molecules in such a way that they retained their thermosensitive properties, including the completely reversible temperature dependence of their fluorescence response. The composite particles developed can be used as local temperature sensors, as carriers for biomolecules, as well as in biosensing and various bioassays employing optical detection schemes.

    ID:1130
  8. Bobrovsky A., Mochalov K., Oleinikov V., Sukhanova A., Prudnikau A., Artemyev M., Shibaev V., Nabiev I. (2012). Optically and electrically controlled circularly polarized emission from cholesteric liquid crystal materials doped with semiconductor quantum dots. Adv. Mater. Weinheim 24 (46), 6216–22 [+]

    Novel types of electro- and photoactive quantum dot-doped cholesteric materials have been engineered. UV-irradiation or electric field application allows one to control the degree of circular polarization and intensity of fluorescence emission by prepared quantum dot-doped liquid crystal films.

    ID:1129
  9. Sukhanova A., EvenDesrumeaux K., Kisserli A., Tabary T., Reveil B., Millot J.M., Chames P., Baty D., Artemyev M., Oleinikov V., Pluot M., Cohen J.H., Nabiev I. (2012). Oriented conjugates of single-domain antibodies and quantum dots: toward a new generation of ultrasmall diagnostic nanoprobes. Nanomedicine 8 (4), 516–25 [+]

    Common strategy for diagnostics with quantum dots (QDs) utilizes the specificity of monoclonal antibodies (mAbs) for targeting. However QD-mAbs conjugates are not always well-suited for this purpose because of their large size. Here, we engineered ultrasmall nanoprobes through oriented conjugation of QDs with 13-kDa single-domain antibodies (sdAbs) derived from llama IgG. Monomeric sdAbs are 12 times smaller than mAbs and demonstrate excellent capacity for refolding. sdAbs were tagged with QDs through an additional cysteine residue integrated within the C terminal of the sdAb. This approach allowed us to develop sdAbs-QD nanoprobes comprising four copies of sdAbs coupled with a QD in a highly oriented manner. sdAbs-QD conjugates specific to carcinoembryonic antigen (CEA) demonstrated excellent specificity of flow cytometry quantitative discrimination of CEA-positive and CEA-negative tumor cells. Moreover, the immunohistochemical labeling of biopsy samples was found to be comparable or even superior to the quality obtained with gold standard protocols of anatomopathology practice. sdAbs-QD-oriented conjugates as developed represent a new generation of ultrasmall diagnostic probes for applications in high-throughput diagnostic platforms.

    ID:1131
  10. Oleinikov V.A. (2009). [Semiconductor fluorescent nanocrystals (quantum dots) in biological biochips]. Bioorg. Khim. 37 (2), 171–89 [+]

    Comprehension of biological processes in cells, tissues and organisms requires identification and analysis of numerous biological objects, mechanisms of their action and regulation. Microarray (biochips) technology is a rare tool to solve this problem. It is based on high-throughput recognition of a target to the probe and has the potential to measure simultaneously the presence of numerous molecules in multiplexed testes, all contained in a small drop of test fluid. Biochips allow the parallel analysis of genomic or proteomic content in healthy versus disease-affected or altered tissues or cells. The signals read-out from the biochips is done with organic dyes which often suffer from photobleaching, low brightness and background fluorescence. Recent data show that the use of fluorescent nanocrystals "quantum dots" (QDs) allows push away these restrictions. The QDs are sufficiently bright to be detected as individual particles, extremely resistant to photobleaching and provide unique possibilities for multiplexing thus supplying the microarray technology with the novel read-out option enabling the sensitivity of detection reaching the single molecule level. This paper is aimed at the development of the approaches to the QDs application in microarray-based detection. Possibilities of QDs application both in solid state (planar) biochips as well as intensively developing technique of suspension biochips (bead-based assays or liquid biochips) are demonstrated. The latter are more and more applied for simultaneous identification of very large numbers of molecules in proteomics, genomics, drug screening and clinical diagnostics. This assays base on spectral encoded elements (as a rule polymer microbeads). The benefits of using optically encoded microbeads (instead of the solid-state two-dimensional arrays) are derived from the freedom of bead to move in three dimensions. Polymeric beads optically encoded with organic dyes allow for a limited number of unique codes, whereas the use of semiconductor nanocrystals as fluorescent tags improves the beads multiplexed imaging capabilities, photostability and sensitivity of the biological objects detection. Additionally, an employment in suspension biochips of Frster resonance energy transfer (FRET) allows improving detection specificity. The absence of fluorescent background from non-interacting with the beads dye-labelled antibodies additionally increases the sensitivity of detection and further facilitates the multiplexing capabilities of nanocrystals-based detection and diagnostics. So the combination of the biochips and QDs techniques allow increasing detection sensitivity and significantly raising the number of detected objects (multiplexing capacities). Such combination should provide the breakthrough in proteomics, particularly in new drugs development, clinical diagnostics, new disease markers identification, better understanding of intracellular mechanisms.

    ID:1132

Vladimir Oleinikov

  • Russia, Moscow, Ul. Miklukho-Maklaya 16/10 — On the map
  • IBCh RAS, build. 34, office. 131
  • Phone: +7(495)335-43-66
  • E-mail: voleinik@mail.ru

Unique scientific setup “System for probe-optical 3D correlative microscopy” (2017-11-28)

Designed, build and included in the register of unique scientific experimental instruments System for probe-optical 3D correlative microscopy”

The new tool is designed to obtain three-dimensional images of biological and other objects, reconstruction of the 3D distribution of various physicochemical parameters along the volume of the object, including the determination of the chemical composition in the local regions of the object with a resolution of the nanometer scale.

All techniques of SPM are available (contact, semicontact, dynamic contact, and tapping modes, force modulation, magnetic and electrostatic force, Kelvin-probe  and scanning spreading resistance microscopy.  Also, it has the possibility to use the confocal fluorescence, Raman, surface and tip enhanced Raman scattering spectroscopy (SERS and TERS), and SNOM. Axial resolution of ultramicrotomography – 20 nm. Instrument contains the hard and software for reconstruction 3-D reconstruction of the morphological/optical structure of samples.

The figure shows the appearance of a unique tool, its scheme and principle of action. As the example of application, the "architecture" of the magnetic-fluorescent microbeads for clinical diagnosis and drug delivery, determined using ultramicrotomography, fluorescent microspectroscopy, and magnetic force microscopy was defined.

Publications

  1. Mochalov K.E., Chistyakov A.A., Solovyeva D.O., Mezin A.V., Oleinikov V.A., Vaskan I.S., Molinari M., Agapov I.I., Nabiev I., Efimov A.E. (2017). An instrumental approach to combining confocal microspectroscopy and 3D scanning probe nanotomography. Ultramicroscopy 182, 118–123 [+]

    In the past decade correlative microscopy, which combines the potentials of different types of high-resolution microscopies with a variety of optical microspectroscopy techniques, has been attracting increasing attention in material science and biological research. One of outstanding solutions in this area is the combination of scanning probe microscopy (SPM), which provides data on not only the topography, but also the spatial distribution of a wide range of physical properties (elasticity, conductivity, etc.), with ultramicrotomy, allowing 3D multiparametric examination of materials. The combination of SPM and ultramicrotomy (scanning probe nanotomography) is very appropriate for characterization of soft multicompound nanostructurized materials, such as polymer matrices and microstructures doped with different types of nanoparticles (magnetic nanoparticles, quantum dots, nanotubes, etc.), and biological materials. A serious problem of this technique is a lack of chemical and optical characterization tools, which may be solved by using optical microspectroscopy. Here, we report the development of an instrumental approach to combining confocal microspectroscopy and 3D scanning probe nanotomography in a single apparatus. This approach retains all the advantages of SPM and upright optical microspectroscopy and allows 3D multiparametric characterization using both techniques. As the first test of the system developed, we have performed correlative characterization of the morphology and the magnetic and fluorescent properties of fluorescent magnetic microspheres doped with a fluorescent dye and magnetic nanoparticles. The results of this study can be used to obtain 3D volume images of a specimen for most high-resolution near-field scanning probe microscopies: SNOM, TERS, AFM-IR, etc. This approach will result in development of unique techniques combining the advantages of SPM (nanoscale morphology and a wide range of physical parameters) and high-resolution optical microspectroscopy (nanoscale chemical mapping and optical properties) and allowing simultaneous 3D measurements.

    ID:1985
  2. Efimov A.E., Agapov I.I., Agapova O.I., Oleinikov V.A., Mezin A.V., Molinari M., Nabiev I., Mochalov K.E. (2017). A novel design of a scanning probe microscope integrated with an ultramicrotome for serial block-face nanotomography. Rev Sci Instrum 88 (2), 023701 [+]

    We present a new concept of a combined scanning probe microscope (SPM)/ultramicrotome apparatus. It enables "slice-and-view" scanning probe nanotomography measurements and 3D reconstruction of the bulk sample nanostructure from series of SPM images after consecutive ultrathin sections. The sample is fixed on a flat XYZ scanning piezostage mounted on the ultramicrotome arm. The SPM measuring head with a cantilever tip and a laser-photodiode tip detection system approaches the sample for SPM measurements of the block-face surface immediately after the ultramicrotome sectioning is performed. The SPM head is moved along guides that are also fixed on the ultramicrotome arm. Thereby, relative dysfunctional displacements of the tip, the sample, and the ultramicrotome knife are minimized. The design of the SPM head enables open frontal optical access to the sample block-face adapted for high-resolution optical lenses for correlative SPM/optical microscopy applications. The new system can be used in a wide range of applications for the study of 3D nanostructures of biological objects, biomaterials, polymer nanocomposites, and nanohybrid materials in various SPM and optical microscopy measuring modes.

    ID:1986