Кафедра лабораторної діагностики біологічних систем

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Detection of air-borne mycotoxin levels by immunobiosensor
(2018) Starodub, Nickolaj; Kluchevich, M.; Székács, A.; Vigera, S.
The main purpose of this article is to investigate the possibility of detection of the level of mycotoxins in air samples by a surface plasmon resonance (SPR) immunobiosensor. Air samples of 0.25-0.5 m3 volume collected using a special miniature air pump device were passed through 20% acetonitrile as an absorption liquid, and 20 μl aliquots of the extract were analyzed by an SPR immunobiosensor. Comparative analyses by the determination of T2 mycotoxin content were carried out in air and in grain samples upon dispersion in a phytotron. Moreover, mycotoxin content was investigated also under field conditions in air and in vegetable matrix, in the absence and presence of Fusarium fungi, and results were shown to have similar characteristics: the level of mycotoxins detected in air samples correlated with that in grain samples. Thus, results of air-borne mycotoxin detection were correlate with mycotoxin levels detected in environmental samples and may used for screening purposes.
Detection of biogenic polyamines in blood of patients with breast cancer
(2019) Prylutskyi, Maksym; Bilko, Nadiia; Starodub, Nickolaj
The main threat of cancer diseases is their spreading throughout the population of many countries of the world and the complexity of their diagnostics at the early stages. Because of that, search and development of the latest diagnostic methods for oncological diseases which would allow them to be diagnosed more precisely and rapidly is going on. The objective of the study was developing a method for qualitative and quantitative analysis of polyamines as potential tumour markers in blood serum of patients with breast cancer using a newly created immune biosensor based on the effect of surface plasmon resonance (SPR). In the process of the study we used basics of immune analysis, methods of biosensor analysis with preliminary modification of biosensor analytical surface with certain reagents, such as protein A and BSA, for better orientation of the sensitive layer made from antibodies. During the study 30 samples of blood serum were analyzed, 21 of which were obtained from the patients with breast cancer and 9 samples were normal, taken from healthy people and used as controls. Analysis of blood serum samples was made, using previously created calibration curve, based on polyamine solutionsin concentrations from 5 ng/mL to 1 μg/mL. It allowed determination of the presence of polyamines in blood samples and approximate concentration of polyamines comparing resonance angle shift in calibration curve and blood samples. According to the obtained results, the concentration of polyamines exceeded their physiological levels and was in the range of 21.3–125.1 ng/mL. The proposed approach allows one to determine the presence and approximate concentrations of polyamines in range from 5 ng/mL to 1 μg/mL in samples of blood serum of patients with breast cancer which correlates with tumour size and the age of the patients.
Detection of polyamines using immunobiosensor based on zinc oxide nanoparticles
(2019) Prylutskyi, Maksym; Starodub, Mykola
Aim: To analyze the performance of biosensor based on nanoparticles of zinc oxide for the detection of spermine and spermidine in solution and in cell culture. Materials and Methods: Zinc oxide nanoparticles were used for preparing biosensor containing antibodies to spermine and spermidine. Polyamine concentration is solutions of spermine and spermidine as well as in lyophilisate of MCF-7 cells was measured by luminescence of the samples excited by laser beam at 380 nm. Results: The minimum concentration for the detection of polyamines in model solutions is 10 ng/ml, and maximum one is 100 ng/ml. A higher level of luminescence intensity of nanoparticles was found during analysis the polyamines in MCF-7 lyophilisate allowing for detecting polyamines at concentrations from 100 cells/ml to 100,000 cells/ml. Conclusions: The proposed biosensor system for determining the level of biogenic polyamines in cell lyophilisate using the optical properties of zinc oxide nanoparticles is promising for further improvement of the methodology and its implementation for detection and measurement of polyamines in biological systems.
Dynamics of Telomere Length and Telomerase Activity in the Human Fetal Liver at 5-12 Weeks of Gestation
(2018) Sorochynska, Khrystyna; Sych, Nataliia; Duda, Alla; Kulebyakina, Kateryna; Krasnienkov, Dmytro; Vaiserman, Alexander; Vatlitsov, Denys
Fetal stem cell- (FSC-) based therapy is a promising treatment option for many diseases. The differentiation potential of FSCs is greater than that in adult stem cells, and they are more tissue-specific and have lower immunogenicity and better intrinsic homing than embryonic ones. Embryonic stem cells have higher proliferative potential than FSCs but can cause teratomas. Therefore, an evaluation of this potential represents an important biomedical challenge. Since regulation of telomere length (TL) is one mechanism governing cellular proliferation, TL is a useful surrogate marker for cell replicative potential. The prenatal dynamics of TL, however, has never been comprehensively studied. In the present study, dynamics of TL and telomerase activity in the human fetal liver during 5–12 weeks of gestation is examined. Both TL and telomerase activity were positively correlated with week of gestation. For both parameters studied, the trend to increase was evident up to 10th week of gestation. After that, they reached a plateau and remained stable. These findings indicate that telomerase activity remains high during the fetal stage, suggesting high replicative capacity of FSCs and their considerable potential for transplantation therapies. These findings, however, are preliminary only due to small sample size and require further evaluation.
Effects of fetal stem cells on life quality of patients with ulcerative colitis
(2018) Demchuk, Mariya; Ivankova, Olena; Klunnyk, Mariya; Matiyashchuk, Iryna; Sych, Nataliia; Sinelnyk, Andriy; Karayev, T.; Shulak, M.; Skalozub, Marina; Sorochynska, Khrystyna
In this article we briefly summarized on the main characteristic features of ulcerative colitis (UC) and accentuated on reduced life quality among the patients with UC, in particular, effect of clinical manifestations of the disease on everyday life was described in such patients. Unambiguously, the principal direction to better life quality in patients with UC consists in decreasing activity of inflammatory process. Significant aspect in solution of this problem for present day suggests use of fetal stem cells (FSCs) in complex treatment of the patients suffering from UC. Specifically, by virtue of stem cell therapy use, rapid lowering of clinical signs of UC was recorded, patients revealed stabilization of hemoglobin and total protein levels in blood. Changes of such parameters in the CG patients tended to be much slower. Within the whole period under study, we recorded a consistent pattern between decreased evidence of underlying disease manifestation that is coupled with higher life quality among the UC patients. In conformity with IBDQ international questionnaire, describing that bowel symptoms, systemic symptoms, emotional and social function, life quality among UC patients of the MG were increased (51.33% vs. 96.43%), whereas the patients of the CG reported the value 61.16% that was higher only within the range of 86.16%. Administration of suspensions with FCSs for complex therapy in UC patients proved to be one of the ways for refining social and psychological wellbeing of the individuals under study.
Influence of human HB-EGF secreted form on cells with different EGFR and ErbB4 quantity
(2019) Krynina, Olga; Korotkevych, Nataliia; Labyntsev, Andrii; Romaniuk, Svitlana; Kolybo, Denys; Komisarenko, Serhii
HB-EGF is one of the most potent ligands of EGFR and ErbB4 receptors. This growth factor plays a pivotal role in many cellular processes, but its effect differs from one cell type to another and remains not fully understood. The aim of this work was to investigate the dependence between the rate of HB-EGF mediated cell proliferation and activation of EGFR and ErbB4 receptors. Therefore, the effects of human recombinant sHB-EGF (rsHB-EGF) on the proliferation of cell lines with different EGFR and ErbB4 quantity and ratio, as well as activation of the MAR K-cascade p38 and ERK1/2 (p42/44) kinases, were analyzed. For comparison, a similar study of the effect of native sHB-EGF secreted by human histiocytic lymphoma cells U937 during co-cultivation with different cell lines was performed. It was proved that cell proliferation in response to sHB-EGF depends not only on the quantity but also on the ratio of EGFR and ErbB4. It was shown that signaling through ErbB4 is associated with activation of p38 kinase and signaling through EGFR associated with activation of ERK1/2 (p42/44) kinase. We assume the existence of two different mechanisms for sHB-EGF-mediated stimulation of cell proliferation, and the simultaneous launch of these mechanisms provides a maximal proliferative response. The results of this study support the feasibility of creating anti-proliferative drugs that target ErbB4.
Ivermectin affects Arabidopsis thaliana microtubules through predicted binding site of β-tubulin
(2024) Kustovskiy, Yevhen; Karpov, Pavel; Blume, Yaroslav; Yemets, Alla
The ivermectin is a potent nematocide and insecticide, which has low toxicity for humans and domestic animals, but due to low biotransformation, it can be dangerous for non-target organisms. The recent determination of ivermectin absorption and accumulation in tissues of higher plants and multiple shreds of evidence of its negative impact on plant physiology provide a basis for the search for ivermectin’s molecular targets and mechanisms of action in plant cells. In this research, for the first time, the ivermectin effect on microtubules of Arabidopsis thaliana cells was studied. It was revealed that ivermectin (250 μg mL- 1) disrupts the microtubule network, induces the loss of microtubule orientation, leads to microtubule curvature and shrinkage, and their longitudinal and cross-linked bundling in various cells of A. thaliana primary roots. Further, the previously proposed binding of ivermectin to the β1-tubulin taxane site was developed and confirmed using molecular dynamics simulations of ivermectin complexes with Haemonchus contortus and A. thaliana β1-tubulins. It was predicted that similar to other microtubule stabilizing agents ivermectin binding causes M-loop stabilization in both H. contortus and A. thaliana β-tubulin, which leads to the enhancement of lateral contacts between subunits of adjacent protofilaments preventing microtubule depolymerization.
New data on histogenesis and histological structure of lung cancer
(2023) Bolgova, L.; Shypko, A.; Tuganova, T.; Alekseenko, O.; Smolanka, I.; Ponomarenko, A.; Bilko, Nadiia
Lung cancer (LC) is one of the most common malignant neoplasms in men around the world, which poses a number of important challenges for scientists. Aim. To analyze the histogenesis, features of the histological structure, and growth of LC. Materials and Methods. The surgical material of 81 patients with LC was studied. Histological preparations were stained with hematoxylin and eosin (H&E) using the Papanicolaou method. Immunohistochemical reactions with monoclonals (Ki-67, PCNA) were conducted. Results. In histological preparations of all LC types (squamous, adenocarcinoma, and small cell), along with solid growth, tumor growth in the alveoli was determined, which started from the basal membrane and grew toward the alveolus center, as evidenced by the morphological features of growth, tumor spread, and development of necrosis in the center. Conclusion. In all the studied histological preparations of LC, tumor growth in the alveoli is noted, which is confirmed by structural and cellular signs and the nature of tumor decay in the alveolus center, which corresponds to the general patterns of development of malignant epithelial tumors.
Optical biosensing analysis of spermine as marker of prostate cancer with help of colloidal gold and anti-spermine antibody modified transducers
(2023) Prylutskyi, Maksym; Zaletok, S.; Khrystosenko, R.; Lebyedyeva, T.; Shpylovyy, P.; Mukha, Yu.; Taran, O.
Prostate cancer (PC) is a deadly oncological disease spread all around the world among elderly men. Existing methods of analysis or detection of traditional biomarkers has proven to be expensive and in some cases less precise. That is why in recent years polyamines are studying as potential PC biomarkers. On the other hand biosensors have already proven themselves as precise and sensitive methods that can be used diagnostics, namely in detection of polyamines. Therefore, the aim of this research was to develop a new type of transducer for SPR biosensor based on use of colloidal gold and compare it with conventional antibody-antigen transducer to detect spermine as prostate cancer biomarker. The analysis was made with help of two biosensing devices Plasmontest and SPR-6 which were used detect spermine in range from 0.5 to 1000 nM During the research it was investigated that transducer treated with colloidal have higher sensivity to spermine comparing with antibody-treated transducer. It was observed that range of detected spermine concentration varies from 0.5 to 1000 nM and oplimum located in range 10-500 nM. This article presents data related to the development of the modern sensitive biosensor capable of detecting spermine as a potential marker of prostate cancer.
Orally administered activated charcoal as a medical countermeasure for acute radiation syndrome in rats
(2021) Snezhkova, Elisaveta; Rodionova, Natalia; Bilko, Denys; Silvestre-Albero, Joaquín; Sydorenko, Alexey; Yurchenko, Olga; Pakharenko, Marharyta; Alavijeh, Mohammad; Bardakhivska, Kvitoslava; Riabchenko, Natalia; Nikolaev, Volodymyr
Activated charcoal (AC) can be taken orally as enterosorbent for treatment of pathological states related to exogenous and endogenous intoxications. Synthesized granulated AC with a highly developed active surface (SBET ~2700 m2/g) was used as a medical countermeasure (MCM) to acute radiation sickness (ARS) in rats after total body X-ray irradiation. AC demonstrates positive results in ARS treatment, as expressed in, (i) a decrease in body weight loss, (ii) a protection of bone marrow (BM) cells colony formation capacity, (iii) a reduction of BM chromosomal aberrations and small intestine and spleen tissue damage, (iv) an amelioration of white blood cell count, and (v) a mitigation of superoxide ion generation rate in the liver. AC oral prescription seems to be perspective modality of ARS treatment.
Peculiarities of sample preparation for the determination of certain mycotoxins in grain products and fruits by immunobiosensor analysis
(2018) Starodub, Nickolaj; Savchuk, M.; Székács, A.; Marty, J.
Our current report focuses on effective ways of sample preparation for detection of mycotoxins aflatoxin B1 and T2 in grain products (wheat, maize, sunflower and oats) as well as patulin in fruits (plum, apple and grape) by the surface plasmon resonance (SPR) immunobiosensor. An effective method for carrying out analysis of mycotoxins by an SPR Immunobiosensor was assessed, and the results obtained were compared with those determined in a traditional ELISA method. The analysis with the immunobiosensor is capable to be carried out in a direct format on the basis of mycotoxin-specific antibodies immobilized on the sensor surface, but its sensitivity was found to be lower than that obtained in a competitive format on the basis of an immobilized mycotoxin conjugate. Due to this characteristic difference in analytical sensitivity, we recommend the former format for screening for the presence of mycotoxins in environmental samples. In turn, the latter format is proposed for verification of the obtained results of analysis. The competitive analysis by the immunobiosensor has demonstrated sensitivity in the range of the traditional ELISA method. Organic solvent extraction of the three mycotoxins, aflatoxin B1, T2 and patulin from produce samples was carried out with acetonitrile and methanol, and the former solvent was concluded that preferable. Particular attention was paid to the problem of matrix components detrimentally affecting extractability and the immunochemical reaction: difficulties due to the presence of small quantities of vegetable oil in grain products needed to be overcome. In cases of samples of substantial vegetable oil content, acetone as an extraction solvent is recommended, the use of which did not appear to affect immunoanalysis results. It was demonstrated that the analysis of patulin in fruits depends on pH of the obtained samples, especially if mycotoxin content is determined in an unripe stage. In such cases, pH determination and neutralization by a special buffer prior to immunoanalytical measurements are recommended.
Role of the heparin-binding domain in intracellular trafficking of sHB-EGF
(2019) Krynina, Olga; Manoilov, Kyrylo; Kolybo, Denys; Komisarenko, Serhii
Heparin-binding EGF-like growth factor (HB-EGF) is a member of the epidermal growth factor family that was proven as a potent mitogen and chemoattractant. HB-EGF mediated EGFR activation is a key event in the stimulation of gene expression, cell migration and proliferation during both normal and pathogenic physiological processes. The main goal of this research was to reveal the role of the heparin-binding domain of HB-EGF in the ligand-receptor formation and its further internalization to the cytoplasm. We used fluorescently-labeled recombinant derivative of soluble HB-EGF and its truncated form (sHB-EGFΔ84–106) with deletion of the heparin-binding domain. Firstly, the binding kinetics of two forms of sHB-EGF to its cell surface receptors was determined using flow cytometry. To determine how the absence of heparin-binding domain in the structure of HB-EGF affects its internalization, we analyzed the endocytosis process of EGFP-sHB-EGFΔ84–106 and EGFP-sHB-EGF complexes by confocal microscopy. It was found that the full-size form of HB-EGF is characterized by a lower intensity of translocation to the cytoplasm in comparison to HBD-deleted form. Thus, differences in the trafficking of the full-size or truncated forms of sHB-EGF in the cell cytoplasm may reflect the mechanisms of extracellular matrix influence on the biological activity of sHB‐EGF.
Study of modifying effects of astaxanthin on cytogenetic manifestations of bystander response in human peripheral blood lymphocytes in vitro
(2021) Pilinska, M.; Shemetun, O.; Talan, O.; Dibska, O.; Talko, V.
To study the possible impact of astaxanthin on the cytogenetic manifestations of the simultaneous development of radiationinduced (RIBE) and tumor-induced bystander effect (TIBE) in human peripheral blood lymphocytes.
Synthesis, properties and bioimaging applications of silver-based quantum dots
(2021) Borovaya, Mariya; Horiunova, Inna; Plokhovska, Svitlana; Pushkarova, Nadia; Blume, Yaroslav; Yemets, Alla
Ag-based quantum dots (QDs) are semiconductor nanomaterials with exclusive electrooptical properties ideally adaptable for various biotechnological, chemical, and medical applications. Silver-based semiconductor nanocrystals have developed rapidly over the past decades. They have become a promising luminescent functional material for in vivo and in vitro fluorescent studies due to their ability to emit at the near-infrared (NIR) wavelength. In this review, we discuss the basic features of Ag-based QDs, the current status of classic (chemical) and novel methods ("green" synthesis) used to produce these QDs. Additionally, the advantages of using such organisms as bacteria, actinomycetes, fungi, algae, and plants for silver-based QDs biosynthesis have been discussed. The application of silver-based QDs as fluorophores for bioimaging application due to their fluorescence intensity, high quantum yield, fluorescent stability, and resistance to photobleaching has also been reviewed.
Treatment Options for Developmental Delay in Children with Perinatal Damage of Nervous System
(2018) Sych, Nataliia; Klunnyk, Mariya; Demchuk, Mariya; Ivankova, Olena; Matiyashchuk, Iryna; Sinelnyk, Andriy; Skalozub, Marina; Sorochynska, Khrystyna
Objective: Improvement of treatment in children with developmental delay (DD) resulting from perinatal affection of CNS using combination with conventional therapy and administration of fetal stem cells (FSCs) in suspensions containing stem cells extracted from human fetal liver and brain. Materials and methods: Our study allocated 14 children diagnosed with developmental delay and perinatal damage of CNS including 4 boys aged 1.5 to 14 years, with their mean age 4.23 ± 0.31 yrs. and 3 girls - from 3.5 to 13 yrs (mean age – 3.92 ± 0.21 yrs.) (Allocated to the MG) where along with conventional therapy were administered fetal stem cells. Control group (CG) included 7 patients with Developmental delay (DD) – children treated with conventional methods only, including 4 males (aged 3 to 14 yrs. and average age – 4.11 ± 0.14 yrs.) and 3 females (aged from 3 to 15 yrs. and mean age 4.02 ± 0.15 yrs.). Results: Our research revealed that fetal stem cell therapy, irrespective of developmental delay severity in children with CNS damage during perinatal period, can favorably influence the course of development and their immunological markers in particular. Procedure of fetal stem cell administration is simple and noninvasive (no potential causes of complications or allergy reactions in the patients). To promote substantial effects of fetal stem cells we additionally recommended a complex treatment with medicines; different supplementary methods were used for the patients. Aspects of fetal stem cells effect in children with developmental delay are not fully understood and do need a thorough scientific research. In particular, future in-depth study is demanded, as well as the larger randomized and placebo-controlled trials to identify a potential of fetal stem cellsrelated advantages in children with developmental delay (DD). Conclusion: Fetal stem cells (FSCs) use in complex treatment of patients with developmental delay tends to improve cognitive functions along with motor development of children with nervous system affection; such patients also revealed increased immunology findings.
Изучение роли протеинкиназ СК1 в организации кортикальных микротрубочек корня Arabidopsis thaliana
(2019) Карпов, П.; Шеремет, Я.; Блюм, Ярослав; Емец, Алла
Представители подсемейства Казеин киназа 1 ( СК1) отличаются межвидовым консерватизмом и обширным набором субстратов фосфорилирования. Недавно появились доказательства того, что СК1 способны непосредственно фосфорилировать тубулин. Таким образом, СК1 вносят вклад в тубулиновый код и функциональную специализацию микротрубочек. В данной работе мы представляем результаты исследования ответной реакции тубулинового цитоскелета растений на обработку СК1 -специфичным ингибитором 04476. Было показано, что D4 4 7 6 оказывает сильный дозозависимый эффект на рост и морфологию корней Arabidopsis thaliana. Эксперименты на растениях, экспресирующих химерный ген gfp-тар4, доказали взаимосвязь наблюдаемых морфологических реакций с пространственной реорганизацией микротрубочек вызванной избирательным ингибированием протеинкиназ СК1.

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