Cell Culture Tests Research Articles

Synthesis and characterization of a chitosan/gelatin transparent film crosslinked with a combination of EDC/NHS for corneal epithelial cell culture scaffold with potential application in cornea implantation

The diseases and disorders of the cornea may lead to blindness, and cornea transplantation has been an effective treatment in this regard. However, lack of cornea throughout the world makes this treatment difficult. Therefore, the first goal of the present study is to make a chitosan/gelatin hyaline film with NHS and EDC crosslinkers for transplanting the epithelial cells of the cornea. Two solutions of gelatin and chitosan were mixed homogeneously before crosslinking in ratios of 20/80, 30/70, 40/60, and 50/50 (Gel/Chi). After 24 hours, they were put in an oven to dry, then EDC and NHS were added to the mixture as crosslinker. Corneal epithelial cell morphology was assessed qualitatively and compared between different ratios of chitosan and gelatin using corneal epithelial cell culture tests. Mechanical and optical analysis were performed; the results implied an increase in chitosan concentration could improve transparency and biocompatibility of produced samples. Fourier Transform Infrared Spectroscopy (FTIR) and SEM analysis were done for the chemical and morphological characterization of fabricated films. Moreover, the presented results revealed that increment in chitosan increased water permeability and degradability of samples. Besides, MTT essays showed that with increment in chitosan concentration, cell proliferation and cell viability were significantly improved.

Calcium Aluminate Cement Blends Containing Bioactive Glass and Strontium for Biomaterial Applications

In this work blends based on calcium aluminate cement (CAC) containing bioactive glass (BG) (5, 7.5 and 10 wt%) and strontium oxide (1 wt%) were produced aiming improve their bioactivity and the capacity to stimulate the bone regeneration. In the first part, the blends containing only BG were characterized as theoretical density, microhardness, uniaxial cold crush strength after SBF and apparent porosity and pore size distribution before and after SBF treatment. In the second part, bioactivity and cell culture tests were also conducted in the blends containing BG and strontium oxide. The addition of 7.5 wt% of BG in homogeneous calcium aluminate cement (CH) improved its mechanical properties as microhardness and uniaxial cold crushing strength. The blends were more bioactive due to the presence of a highly soluble amorphous phase as confirmed by means of SEM/EDX mainly for 7.5 wt% BG without and with Sr from 1 day in SBF. FTIR analyses indicated the formation the apatite-like phase by means of increase of intensity of the PO43- peaks after SBF treatment. All blends allowed the development of the osteoblastic phenotype and the formation of mineralized matrix increased due to the inclusion of BG and Sr promoting the osteogenesis process.

Controllable design of a nano-bio aptasensing interface based on tetrahedral framework nucleic acids in an integrated microfluidic platform

The limited reaction time and sample volume in the confined space of microfluidic devices give considerable importance to the development of more effective biosensing interfaces. Herein, the self-assembling of tetrahedral framework nucleic acids (FNAs) with controllable size on the interface of the microfluidic microchannels is studied. Compared with macroscopic turbulence control on traditional micro-structured microfluidic surface, the novel FNA-engineered microfluidic interface successfully constructs a 3D reaction space at nanoscale by raising DNA probes away from the surface. This FNA interface dramatically improves the reaction kinetics during molecular recognition due to extremely ordered orientation, configuration and density of DNA probes on the surface. Finally, the FNA-engineered interface is applied in a novel multi-functional microfluidic platform, towards a “one-stop” assay of Escherichia coli O157: H7 (E. coli O157: H7), integrating capture, release, enrichment, cell culture and antimicrobial susceptibility testing (AST). With the FNA-aptamer probe, we achieved an enhanced bacterial detecting efficiency (10 CFU/mL) plus excellent selectivity and precision. The appicability was strongly demonstrated when the biosensor was successfully applied in real samples, including the analysis of antibiotic susceptibility and minimum inhibitory concentration (MIC) of E. coli O157: H7 among different antibiotics. The application of FNA interface will open a wide avenue for the development of microfluidic biosensors for other pathogenic microorganisms or circulating tumor cells (CTC) simply by changing the aptamers.

An in-vitro study: The effect of surface properties on bioactivity of the oxide layer fabricated on Zr substrate by PEO

Surface properties can affect cell attachment and growth ability on the biomaterials. In this work, the effect of the morphology and chemical composition of the surface coatings synthesized on zirconium (Zr) substrates on bioactivity was studied. The surface of pure Zr was modified via a plasma electrolytic oxidation process to obtain oxide-based coatings with different surface chemical compositions and pore sizes by changing the process parameters in the same electrolyte. The characterization of the coatings was conducted using X-ray diffraction (XRD), scanning electron microscopy (SEM), contact angle goniometry (CAG) and energy dispersive spectroscopy (EDS), and Fourier transform infrared spectroscopy (FTIR). The results showed that the coatings comprised t-ZrO2 and Ca0.134Zr0.86O1.86 phases. During the oxidation of Zr substrates, applying an anode current with different densities and frequencies altered the surface roughness Ra of the samples. The relationship between the surface morphology, chemical composition, and bioproperties was investigated by simulated body fluid (SBF) and cell culture tests. The coated samples were immersed into SBF solution and the formation of hydroxyapatite (HA) structure on the surface was detected after 7 days of immersion. A close relationship between the development of HA, the size of the porosity and chemical composition of the coating surface was observed. Additionally, the cell attachment and cell growth on the surface were found to be dependent on the chemistry and topography of coated surfaces. The cell culture test revealed that the growth of the cells was improved with increasing pore sizes on the coating surface.

Proliferation of mesenchymal stem cells by graphene-attached soft material structure

This paper aims to clarify the effect of a structure of graphene attached on soft material, as against conventional rigid materials, and the thereby-obtained graphene-based micro/nano-scale surface structures on proliferation of human mesenchymal stem cells (MSCs). The cell culture tests are conducted using graphene-attached poly(dimethylsiloxane) (PDMS) substrates having various surface structures and graphene-attached glass substrates as the scaffolds. The fluorescence microscopy observation results demonstrate that the existence of graphene film on PDMS significantly promotes the cellular proliferation and elongation in both cases using two kinds of media, and the rate of cell growth for the graphene-attached PDMS substrates is much larger compared to the graphene-attached glass substrates. The difference in the graphene-based surface structures on PDMS has little influence on the degree of cell growth. It is implied that the number of cells corresponds to the rate of graphene covering the surface of PDMS, suggesting that the cell growth strongly depends on the projected surface area of the graphene film attached on PDMS.

Fabrication of bioactive novel scaffold by solid‐state reaction sintering for bone regeneration

In this work we present a new type of scaffold obtained by solid-state reaction, simultaneous sintering of a mixture of precursor oxides, carbonates, and organic substances, the latter used for pore generation. Having variable local composition, exhibits excellent overall physicochemical and bioactivity response. Open porosity is about 50%-60% and its permeability 10-11 m2 . X-ray diffraction exhibits the presence of a sodium-calcium silicate and sodium-calcium phosphate crystalline phases. Additionally, by mechanical compression tests the range of failure stress obtained for the scaffolds was 0.3-1.1MPa. The bioactivity and dissolution rate of the scaffolds were evaluated by in vitro tests. After 1week soaking in simulated body fluid, the formation of a continuous hydroxyapatite layer, which does not differentiate local compositions, was observed. Our results from cell culture tests clearly indicate that during hydroxyapatite layer formation, scaffolds do not liberate any cytotoxic substances. Moreover, cells seeded in the hydroxyapatite-covered scaffolds grew better than the control.

Evaluation of a SARS-CoV-2 rapid antigen test: Potential to help reduce community spread?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can spread from symptomatic patients with COVID-19, but also from asymptomatic individuals. Therefore, robust surveillance and timely interventions are essential for the control of virus spread within the community. In this regard the frequency of testing and speed of reporting, but not the test sensitivity alone, play a crucial role. In order to reduce the costs and meet the expanding demands in real-time RT-PCR (rRT-PCR) testing for SARS-CoV-2, complementary assays, such as rapid antigen tests, have been developed. Rigorous analysis under varying conditions is required to assess the clinical performance of these tests and to ensure reproducible results. We evaluated the sensitivity and specificity of a recently licensed rapid antigen test using 137 clinical samples in two institutions. Test sensitivity was between 88.2-89.6 % when applied to samples with viral loads typically seen in infectious patients. Of 32 rRT-PCR positive samples, 19 demonstrated infectivity in cell culture, and 84 % of these samples were reactive with the antigen test. Seven full-genome sequenced SARS-CoV-2 isolates and SARS-CoV-1 were detected with this antigen test, with no cross-reactivity against other common respiratory viruses. Numerous antigen tests are available for SARS-CoV-2 testing and their performance to detect infectious individuals may vary. Head-to-head comparison along with cell culture testing for infectivity may prove useful to identify better performing antigen tests. The antigen test analyzed in this study is easy-to-use, inexpensive, and scalable. It can be helpful in monitoring infection trends and thus has potential to reduce transmission.

Fabrication and characterization of Ag- and Ga-doped mesoporous glass-coated scaffolds based on natural marine sponges with improved mechanical properties.

Natural marine sponges were used as sacrificial template for the fabrication of bioactive glass-based scaffolds. After sintering at 1050°C, the resulting samples were additionally coated with a silicate solution containing biologically active ions (Ag and Ga), well-known for their antibacterial properties. The produced scaffolds were characterized by superior mechanical properties (maximum compressive strength of 4MPa) and total porosity of ~80% in comparison to standard scaffolds made by using PU foam templates. Direct cell culture tests performed on the uncoated and coated samples showed positive results in terms of adhesion, proliferation, and differentiation of MC3T3-E1 cells. Moreover, vascular endothelial growth factor (VEGF) secretion from cells in contact with scaffold dissolution products was measured after 7 and 10 days of incubation, showing promising angiogenic results for bone tissue engineering applications. The antibacterial potential of the produced samples was assessed by performing agar diffusion tests against both Gram-positive and Gram-negative bacteria.

A locust embryo as predictive developmental neurotoxicity testing system for pioneer axon pathway formation.

Exposure to environmental chemicals during in utero and early postnatal development can cause a wide range of neurological defects. Since current guidelines for identifying developmental neurotoxic chemicals depend on the use of large numbers of rodents in animal experiments, it has been proposed to design rapid and cost-efficient in vitro screening test batteries that are mainly based on mixed neuronal/glial cultures. However, cell culture tests do not assay correct wiring of neuronal circuits. The establishment of precise anatomical connectivity is a key event in the development of a functional brain. Here, we expose intact embryos of the locust (Locusta migratoria) in serum-free culture to test chemicals and visualize correct navigation of identified pioneer axons by fluorescence microscopy. We define separate toxicological endpoints for axonal elongation and navigation along a stereotyped pathway. To distinguish developmental neurotoxicity (DNT) from general toxicity, we quantify defects in axonal elongation and navigation in concentration-response curves and compare it to the biochemically determined viability of the embryo. The investigation of a panel of recognized DNT-positive and -negative test compounds supports a rather high predictability of this invertebrate embryo assay. Similar to the semaphorin-mediated guidance of neurites in mammalian cortex, correct axonal navigation of the locust pioneer axons relies on steering cues from members of this family of cell recognition molecules. Due to the evolutionary conserved mechanisms of neurite guidance, we suggest that our pioneer axon paradigm might provide mechanistically relevant information on the DNT potential of chemical agents on the processes of axon elongation, navigation, and fasciculation.

Effects of the sources of calcium and phosphorus on the structural and functional properties of ceramic coatings on titanium dental implants produced by plasma electrolytic oxidation

Plasma Electrolytic Oxidation (PEO) is as a promising technique to modify metal surfaces by application of oxide ceramic coatings with appropriate physical, chemical and biological characteristics. Therefore, objective of this research was to find the simplest settings, yet able to produce relevant bioactive implant surfaces layers on Ti implants by means of PEO. We show that an electrolyte containing potassium dihydrogen phosphate as a source of P and either calcium hydroxide or calcium formate as a source of Ca in combination with a chelating agent, ethylenediamine tetraacetic acid (EDTA), is suitable for PEO to deliver coatings with desired properties. We determined surface morphology, roughness, wettability, chemical and phase composition of titanium after the PEO process. To investigate biocompatibility and bacterial properties of the PEO oxide coatings we used microbial and cell culture tests. The electrolyte based on Ca(OH)2 and EDTA promotes active crystallization of apatites after PEO processing of the Ti implants. The PEO layers can increase electrochemical corrosion resistance. The PEO can be potentially used for development of bioactive surfaces with increased support of eukaryotic cells while inhibiting attachment and growth of bacteria without use of antibacterial agents.

The Neat Dance of COVID-19: NEAT1, DANCR, and Co-Modulated Cholinergic RNAs Link to Inflammation.

The COVID-19 pandemic exerts inflammation-related parasympathetic complications and post-infection manifestations with major inter-individual variability. To seek the corresponding transcriptomic origins for the impact of COVID-19 infection and its aftermath consequences, we sought the relevance of long and short non-coding RNAs (ncRNAs) for susceptibility to COVID-19 infection. We selected inflammation-prone men and women of diverse ages among the cohort of Genome Tissue expression (GTEx) by mining RNA-seq datasets from their lung, and blood tissues, followed by quantitative qRT-PCR, bioinformatics-based network analyses and thorough statistics compared to brain cell culture and infection tests with COVID-19 and H1N1 viruses. In lung tissues from 57 inflammation-prone, but not other GTEx donors, we discovered sharp declines of the lung pathology-associated ncRNA DANCR and the nuclear paraspeckles forming neuroprotective ncRNA NEAT1. Accompanying increases in the acetylcholine-regulating transcripts capable of controlling inflammation co-appeared in SARS-CoV-2 infected but not H1N1 influenza infected lung cells. The lung cells-characteristic DANCR and NEAT1 association with inflammation-controlling transcripts could not be observed in blood cells, weakened with age and presented sex-dependent links in GTEx lung RNA-seq dataset. Supporting active involvement in the inflammatory risks accompanying COVID-19, DANCR’s decline associated with decrease of the COVID-19-related cellular transcript ACE2 and with sex-related increases in coding transcripts potentiating acetylcholine signaling. Furthermore, transcription factors (TFs) in lung, brain and cultured infected cells created networks with the candidate transcripts, indicating tissue-specific expression patterns. Supporting links of post-infection inflammatory and cognitive damages with cholinergic mal-functioning, man and woman-originated cultured cholinergic neurons presented differentiation-related increases of DANCR and NEAT1 targeting microRNAs. Briefly, changes in ncRNAs and TFs from inflammation-prone human lung tissues, SARS-CoV-2-infected lung cells and man and woman-derived differentiated cholinergic neurons reflected the inflammatory pathobiology related to COVID-19. By shifting ncRNA differences into comparative diagnostic and therapeutic profiles, our RNA-sequencing based Resource can identify ncRNA regulating candidates for COVID-19 and its associated immediate and predicted long-term inflammation and neurological complications, and sex-related therapeutics thereof. Our findings encourage diagnostics of involved tissue, and further investigation of NEAT1-inducing statins and anti-cholinergic medications in the COVID-19 context.

Formation and cytocompatibility of a hierarchical porous coating on Ti-20Zr-10Nb-4Ta alloy by micro-arc oxidation

Beta-Ti alloy has great potential for hard tissue replacement in the biomedical field. Here, a novel hierarchical porous coating was directly prepared on a beta-Ti alloy of Ti-20Zr-10Nb-4Ta by micro-arc oxidation (MAO) in a Na2B4O7 electrolyte for improving its bioactivity. The microstructure, phase composition, adhesion to the substrate, hydrophilicity, and cytocompatibility of the coatings were evaluated, and the effect of the interaction between alloying elements and the electrolyte on the coating formation was discussed. The hierarchical structure of the coating consisted of submicron pores and layered micron pores. It was mainly composed of rutile TiO2 and a small amount of oxides formed by alloying elements. The adhesion to the substrate was improved due to the lower internal stress. The coating exhibited excellent hydrophilicity (water contact angle: 10.6 ± 1.6°). The analysis of the coating morphology evolution showed that the oxide ejected from the discharge channel during MAO was partially dissolved by the Na2B4O7 electrolyte, which was the main reason for the formation of the hierarchical porous structure. The cell culture tests demonstrated that the novel coating had better cell adhesion and spreading ability, which indicates that this coating on the beta-Ti alloy may have significant potential in dental and orthopedic applications.

Preparation of phospholipid-based polycarbonate urethanes for potential applications of blood-contacting implants.

Polyurethanes are widely used in interventional devices due to the excellent physicochemical property. However, non-specific adhesion and severe inflammatory response of ordinary polyurethanes may lead to severe complications of intravenous devices. Herein, a novel phospholipid-based polycarbonate urethanes (PCUs) were developed via two-step solution polymerization by direct synthesis based on functional raw materials. Furthermore, PCUs were coated on biomedical metal sheets to construct biomimetic anti-fouling surface. The results of stress–strain curves exhibited excellent tensile properties of PCUs films. Differential scanning calorimetry results indicated that the microphase separation of such PCUs polymers could be well regulated by adjusting the formulation of chain extender, leading to different biological response. In vitro blood compatibility tests including bovine serum albumin adsorption, fibrinogen adsorption and denaturation, platelet adhesion and whole-blood experiment showed superior performance in inhibition non-specific adhesion of PCUs samples. Endothelial cells and smooth muscle cells culture tests further revealed a good anti-cell adhesion ability. Finally, animal experiments including ex vivo blood circulation and subcutaneous inflammation animal experiments indicated a strong ability in anti-thrombosis and histocompatibility. These results high light the strong anti-adhesion property of phospholipid-based PCUs films, which may be applied to the blood-contacting implants such as intravenous catheter or antithrombotic surface in the future.

Ready to go 3D? A semi-automated protocol for microwell spheroid arrays to increase scalability and throughput of 3D cell culture testing

3-dimensional (3D) cell cultures are being increasingly recognized as physiologically more relevant in vitro models than traditional monolayer cultures, because they better mimic in vivo-like microenvironment, cell-cell and cell-extracellular matrix interactions. Nevertheless, the broader use of 3D models might be limited by requirements for special consumables, equipment, or skills for 3D cell cultures, and by their limited throughput and scalability. In this study, we optimized and adapted a commercially available agarose-micromolding technique to produce scaffold-free spheroid cultures. Brightfield microscopy was used for routine nondestructive and noninvasive evaluation of spheroid formation and growth. The workflow is compatible with manual, as well as high speed automated microscopic image acquisition, and it is supplemented with an in-house developed macro ‘Spheroid_Finder’ for open source software Fiji to facilitate rapid automated image analysis. This protocol was used to characterize and quantify spheroid formation and growth of two different hepatic cell lines, hTERT immortalized, but non-cancerous, adult human liver stem cell line HL1-hT1, and human hepatocellular carcinoma cell line HepG2, as well as their responses to a model antiproliferative and cytotoxic agent, 5-fluorouracil. The complete protocol provides a simple and ready-to-use solution to initiate scaffold-free spheroid cultures in any laboratory with standard equipment for mammalian in vitro cell culture work. Thus, it allows to increase throughput and scale of spheroid culture experiments, which can be greatly utilized in different areas of biomedical, pharmaceutical and toxicological research.

Polydopamine (PDA) mediated nanogranular-structured titanium dioxide (TiO2) coating on polyetheretherketone (PEEK) for oral and maxillofacial implants application

Polyetheretherketone (PEEK) is being used in various biomedical devices thanks to its bio-inertness, nontoxicity, and an elastic modulus near to that of human cortical bone. However, its oral and maxillofacial applications remain challenging owing to is hydrophobicity, lack of biocompatibility, and infectious complications. To address this problem, a biocompatible nanogranular-structured TiO2 coating assisted by a mediating layer of pre-grafted polydopamine (PDA) was deposited onto PEEK to modify its surface. The PDA-mediated TiO2 coated PEEK showed a unique surface layer of single anatase phase. The surface tended to become more hydrophilic with increased TiO2 deposition, and even turned super-hydrophilic with a sufficient amount of deposited TiO2. In vitro cell culture tests verified that such prepared PEEK improved adhesion and proliferation of both fibroblast and osteoblast cells, decreased the inflammatory reaction by suppressing TNF-α expression for macrophages, as well as enhanced the antibacterial activity against Staphylococcus aureus and Streptococcus mutans bacteria attachment when compared to only PDA coated and uncoated PEEK. Such a superior bio-performance is apparently correlated with anatase TiO2, as well as its improved physiochemical surface states, including altered surface energy. The PDA-mediated TiO2 nanostructured coating may bode well for oral and maxillofacial implants applications of PEEK.

IGF-1 loaded injectable microspheres for potential repair of the infarcted myocardium.

The use of injectable scaffolds to repair the infarcted heart is receiving great interest. Thermosensitive polymers, in situ polymerization, in situ cross-linking, and self-assembling peptides are the most investigated approaches to obtain injectability.Aim of the present work was the preparation and characterization of a novel bioactive scaffold, in form of injectable microspheres, for cardiac repair. Gellan/gelatin microspheres were prepared by a water-in-oil emulsion and loaded by adsorption with Insulin-like growth factor 1 to promote tissue regeneration. Obtained microspheres underwent morphological, physicochemical and biological characterization, including cell culture tests in static and dynamic conditions and in vivo tests. Morphological analysis of the microspheres showed a spherical shape, a microporous surface and an average diameter of 66 ± 17µm (under dry conditions) and 123 ± 24 µm (under wet conditions). Chemical Imaging analysis pointed out a homogeneous distribution of gellan, gelatin and Insulin-like growth factor-1 within the microsphere matrix. In vitro cell culture tests showed that the microspheres promoted rat cardiac progenitor cells adhesion, and cluster formation. After dynamic suspension culture within an impeller-free bioreactor, cells still adhered to microspheres, spreading their cytoplasm over microsphere surface. Intramyocardial administration of microspheres in a cryoinjury rat model attenuated chamber dilatation, myocardial damage and fibrosis and improved cell homing.Overall, the findings of this study confirm that the produced microspheres display morphological, physicochemical, functional and biological properties potentially adequate for future applications as injectable scaffold for cardiac tissue engineering.

In vitro study of bioactive glass coatings obtained by atmospheric plasma spraying

This research has addressed a complete study of the bioactivity of bioactive glass coatings obtained by atmospheric plasma spraying. The coatings have been characterized in terms of microstructure, adhesion, crystalline phases and bioactivity. Hydroxycarbonate apatite formation was also monitored following a standard protocol and the in vitro cell response was evaluated by human osteoblast-like cells (MG-63 cells) incubation.The obtained coatings shown a microstructure typical of glass coatings. A simulated body fluid test proved that coatings are capable of developing a surface layer of hydroxycarbonate apatite whereas the appearance of this phase takes place at a longer time than that observed for the powder feedstock. Cell-culture test showed multidirectional growth of MG-63 cells which promoted good contact between cells and the surface of the coating. This study has confirmed a positive effect of the coatings in terms of surface bioactivity and, more interestingly, it has proven an adequate cell-material interaction on the coating surface.

Immunoconjugates to increase photoinactivation of bovine alphaherpesvirus 1 in semen

Artificial insemination and in vitro embryo production are increasingly used to improve the reproductive efficiency of herds, however success of these techniques depends on the sanitary quality of the semen. Insemination centers commonly use antibiotics in their routine procedure, but they are not able against viruses. In this paper, we demonstrate a new approach for disinfecting virus in bovine semen using photoimmunoinactivation, an adaptation of the photodynamic inactivation (PDI) methodology. The photosensitizers (PSs), hematoporphyrin (HP) and zinc tetracarboxy-phthalocyanine (ZnPc) were conjugated to Immunoglobulin Y (IgY) anti-bovine alphaherpesvirus 1 (BoHV-1) and used for PDI against the BoHV-1 viruses in cell culture and compared to the unconjugated PSs. Both treatments proved to be efficient, but a significant decrease in the irradiation time required to completely eliminate the virus was observed in the samples treated with the immunoconjugates. Photophysical measurements help us to understand the coupling between PSs and IgY and the evaluated production of singlet oxygen. Following the cell culture test, the same approach was applied in semen artificially infected with BoHV-1. The immunoconjugates were also efficient for complete virus inactivation up to 5 min of irradiation and proved to be safe using several parameters of sperm viability, demonstrating the feasibility of our strategy for disinfection viruses in semen.

Rapid and highly effective bacteria-killing by polydopamine/IR780@MnO2–Ti using near-infrared light

Bacterial infections are becoming a serious public health challenge because the abuse of antibiotics worldwide has caused bacterial resistance and the occurrence of superbugs. In this work, a new near-infrared (NIR) photoresponsive coating was constructed on Ti implant material, which was composed of MnO2, IR780 photosensitizer and polydopamine (PDA). Under the irradiation of 808 nm NIR light, the component of MnO2 in the coating exhibited high photothermal performance, and IR780 could be stimulated to generate ROS as a photosensitizer. The addition of PDA not only improved the cytocompatibility of the composite coating but also strengthened the light absorption ability of the system. Under the irradiation of 808 nm NIR light for 15 min, the composite coating killed 99.94% Staphylococcus aureus and 99.89% Escherichia coli on Ti plates. This rapid and highly effective bactericidal rate was ascribed to the synergistic effects of photothermal performance and photodynamic effect of the coatings excited by NIR light. Cell culture tests showed that PDA/IR780@MnO2–Ti had good cytocompatibility due to the excellent biocompatibility of PDA and nanostructured surface. Therefore, this surface system will be a very promising plateform to safely and effectively kill bacteria on biomedical devices and implants in a short time.

Development and use of indirect liquid-phase ELISA test system for detection of PRRS virus antigen during in-process control of raw materials intended for vaccine production

Porcine reproductive and respiratory syndrome (PRRS) being endemic and reported in the most countries in the world remains one of the most challenging diseases in pig industry. The main disease control measures include preventive vaccination and animal movement control within and outside the country as well as diagnostic testing of pigs in the population. Live and inactivated vaccines are used for specific prevention of porcine reproductive and respiratory syndrome. Complete and irreversible infectious agent inactivation with maximum epitope preservation and protective immunity in immunized animals are the main requirements for inactivated vaccines. Therefore, continuous improvement of methods for vaccine quality control at various vaccine production stages is of current importance. Results of development of the test system based on indirect liquid-phase enzyme-linked immunosorbent assay (ELISA) for PRRS virus antigen detection and activity testing in infectious and inactivated virus-containing cell cultures at intermediate stages of the vaccine production process are described in the paper. The test-system development process included purified and concentrated virus antigen as well as hyperimmune rabbit sera preparation. Specificity of purified and concentrated virus antigen was confirmed with real-time polymerase chain reaction. The developed test-system was shown to detect the virus antigen at initial infectivity titre of 4.87–7.21 lg TCID50/cm3 corresponding to ELISA titre (dilution) of 1:4 up to 1:64. Methodical Guidelines for detection of porcine reproductive and respiratory syndrome virus antigen with indirect liquid-phase enzyme-linked immunosorbent assay (ELISA) (2019) were developed based on the work results, commissioned and approved by the FGBI “ARRIAH” Scientific Board.