Here, we propose and study several types of quartz surface coatings designed for the high-performance sorption of biomolecules and their subsequent detection by a photonic crystal surface mode (PC ...SM) biosensor. The deposition and sorption of biomolecules are revealed by analyzing changes in the propagation parameters of optical modes on the surface of a photonic crystal (PC). The method makes it possible to measure molecular and cellular affinity interactions in real time by independently recording the values of the angle of total internal reflection and the angle of excitation of the surface wave on the surface of the PC. A series of dextrans with various anchor groups (aldehyde, carboxy, epoxy) suitable for binding with bioligands have been studied. We have carried out comparative experiments with dextrans with other molecular weights. The results confirmed that dextran with a Mw of 500 kDa and anchor epoxy groups have a promising potential as a matrix for the detection of proteins in optical biosensors. The proposed approach would make it possible to enhance the sensitivity of the PC SM biosensor and also permit studying the binding process of low molecular weight molecules in real time.
We propose and demonstrate dendrimer-based coatings for a sensitive biochip surface that enhance the high-performance sorption of small molecules (i.e., biomolecules with low molecular weights) and ...the sensitivity of a label-free, real-time photonic crystal surface mode (PC SM) biosensor. Biomolecule sorption is detected by measuring changes in the parameters of optical modes on the surface of a photonic crystal (PC). We describe the step-by-step biochip fabrication process. Using oligonucleotides as small molecules and PC SM visualization in a microfluidic mode, we show that the PAMAM (poly-amidoamine)-modified chip's sorption efficiency is almost 14 times higher than that of the planar aminosilane layer and 5 times higher than the 3D epoxy-dextran matrix. The results obtained demonstrate a promising direction for further development of the dendrimer-based PC SM sensor method as an advanced label-free microfluidic tool for detecting biomolecule interactions. Current label-free methods for small biomolecule detection, such as surface plasmon resonance (SPR), have a detection limit down to pM. In this work, we achieved for a PC SM biosensor a Limit of Quantitation of up to 70 fM, which is comparable with the best label-using methods without their inherent disadvantages, such as changes in molecular activity caused by labeling.
Background: The pandemic of the new coronavirus infection, COVID-19, is currently ongoing in the world. Over the years, the pathogen, SARS-CoV-2, has undergone a series of mutational genome changes, ...which has led to the spread of various genetic variants of the virus. Meanwhile, the methods used to diagnose SARS-CoV-2, to establish the disease stage and to assess the immunity, are nonspecific to SARS-CoV-2 variants and time-consumable. Thus, the development of new methods for diagnosing COVID-19, as well as their implementation in practice, is currently an important direction. In particular, application of systems based on chemically modified fluorescent microspheres (with a multiplex assay for target protein molecules) opens great opportunities. Aim: development of a microfluidic diagnostic test system based on fluorescent microspheres for the specific detection of immunoglobulins G (IgG) to SARS-CoV-2. Methods: A collection of human serum samples was characterized using enzyme-linked immunosorbent assay (ELISA) and commercially available reagent kits. IgG to SARS-CoV-2 in the human serum were detected by the developed immunofluorescent method using microspheres containing the chemically immobilized RBD fragment of the SARS-CoV-2 (Kappa variant) viral S-protein. Results: The level of IgG in the blood serum of recovered volunteers was 9-300 times higher than that in apparently healthy volunteers, according to ELISA (p0.001). Conjugates of fluorescent microspheres with the RBD-fragment of the S-protein, capable of specifically binding IgG from the blood serum, have been obtained. The immune complexes formation was confirmed by the fluorescence microscopy data; the fluorescence intensity of secondary antibodies in the immune complexes formed on the surface of microspheres was proportional to the content of IgG (r 0.963). The test system had a good predictive value (AUC 70.3%). Conclusion: A test system has been developed, based on fluorescent microspheres containing the immobilized RBD fragment of the SARS-CoV-2 S-protein, for the immunofluorescent detection of IgG in the human blood serum. When testing the system on samples with different levels of IgG to SARS-CoV-2, its prognostic value was shown. The obtained results allow us to present the test system as a method to assess the level of immunoglobulins to SARS-CoV-2 in the human blood serum for the implementation in clinical practice. The test system can also be integrated into various microfluidic systems to create chips and devices for the point-of-care diagnostics.
Background. Despite on the general trend towards decreasing the incidence of newly diagnosed active forms of tuberculosis, the situation with spreading of this disease in Russian Federation remains ...extremely tense. At the same time, the diagnosis is carried out according to the standard scheme, which takes about a month; another month takes test formulation for drug sensitivity. Thus, the development of new methods for diagnostics and typing of mycobacteria, as well as practice implementation of these developments is an urgent direction. Modern developments in the field of microfluidic technologies open up great opportunities in this direction. Aim. Development of a method for identification and typing of Mycobacterium tuberculosis using a label-free biosensor on surface waves in a one-dimensional photonic crystal (PC SM biosensor). Methods. Oligonucleotide probes were selected and synthesized as DNA targets for M. tuberculosis typing. The photonic crystal surface was modified with aqueous solutions of (3-aminopropyl)triethoxysilane, Leuconostoc mesenteroides dextrans and bovine serum albumin. Experiments were carried out using a PC SM biosensor. Results. Sequences of detecting oligonucleotide probes were selected for spoligotyping of M. tuberculosis on the PC SM biosensor. Modification of their 3'-ends was carried out in order to create extended single-stranded regions that are not subject to the formation of secondary structures and facilitate hybridization with a single-stranded DNA target. Several series of experimental modifications of the PC surface were carried out by using L. mesenteroides dextrans with different functional groups (including detection of the modification results real time) with simultaneous registration of the increment layer size and volume refractive index of the mixture, which excludes the use of a reference cell. Other experiments were carried out to detect the specific binding of biotinylated oligonucleotide probes to the modified PC surface. Conclusions. A technique for the design of probes was developed and a model system of oligonucleotides for the detection of single-stranded DNA using a PC biosensor was proposed. The developed technique of modification of the PC surface with dextrans from L. mesenteroides, which allows to increase the sensitivity of detection of oligonucleotides using the PC SM biosensor. This approach will further expand the panel of diagnostic probes, including identification of resistance markers.
Immunotropic drugs (such as sera, blood plasma and immunoglobulins) have polyfunctional activity with anti-infectious, antitoxic, homeostasis-regulating properties. Based on this, we conducted the ...present study with the purpose to evaluate the therapeutic effect of anti-cadmium -anti-radiation serum (AARS) on the level of radiation-chemical damage. Modeling of experimental radiation-chemical damage was carried out by preliminary 4-time subcutaneous injection of cadmium chloride solution (CdCl
2
) at doses of 1/500 LD
50
, followed by radiation exposure of 90 outbred white rats at doses of 7.0 and 9.0 Gy (LD
50
and LD
100
, respectively). It was found that a single subcutaneous injection of TDC at a dose of 25 mg/kg of live weight 24 hours after the combined radiation-toxic damage had a radioprotective and antitoxic effect, increasing the survival rate of animals affected by lethal doses of CdCl
2
and γ-rays. Increase in the survival rate of animals treated with the test agent after exposure to two-factor damage was due to inhibition of the concentration of BFR-active toxic compounds and reciprocal increase in the synthesis of metallothioneins and SOD. Thus, the use of immunotropic agent of anti-cadmium anti-radiation serum (AARS) after exposure to two-factor combined radiation-chemical damage provided 90% survival rate by induction of increased formation of metallothioneins and SOD, inhibition of redox-cycled toxic radicals formation and neutralization of cadmium toxicity with the help of anti-cadmium antibodies.
Purpose of the study. Study of the nearest results of laparoscopic operations in patients with localized renal cell carcinoma.Materials and methods. The results of treatment of 92 patients who ...underwent operative treatment in the period from 2015 to 2017 with the help of laparoscopic technique for kidney carcinoma were analyzed and evaluated. Of these, 57 men (62%) and 35 women (38%), the average age is 50.5 ± 2.25 years (min. 26 and not more than 75 years). Results. The duration of the operation was influenced by various factors, such as the features of the vascular anatomy of the kidneys, the presence of the adhesive process of the abdominal cavity, the state of paranephalic fat. When improving the technique of performing laparoscopic interventions, the operation time was reduced. In the period of mastering the technique of laparoscopic interventions on the kidney, the operation time averaged 210.0 ± 20.2 min (min 120 and max 300 min); subsequently it decreased on average to 130.00 ± 10.5 min (min 70 and max 190 min). After accumulation of experience, the technique of laparoscopic resection of the kidney without total ischemia with selective clamping of segmental arteries, or without compression of the renal artery was mastered.The conclusion. An analysis of the immediate results of laparoscopic interventions in patients with renal cell carcinoma showed their high efficacy and safety. The results of laparoscopic nephrectomies and kidney resections significantly improve with the improvement of technology and the accumulation of surgical experience.
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