Carbodiimide Method Research Articles

Disulfiram-Loaded PLGA nanoparticles modified with a Phenyl borate chitosan Conjugate enhance hepatic carcinoma treatment.

Disulfiram (DSF), which has been traditionally used to treat alcoholism, has been shown to inhibit tumor growth, indicating its potential as an anticancer agent. However, its development and application are hindered by its poor water solubility, instability in physiological environments, and low bioavailability. In this study, phenylboronic acid-chitosan (PBA-CS) grafts were synthesized using the carbodiimide method. PBA-CS-modified DSF PLGA nanoparticles (DSF@PBA-CS-PLGA NPs) were constructed by coating the nanoparticle surfaces with PBA-CS to improve the stability of DSF in physiological environments and enhance its anti-tumor effects. The structures of PBA-CS and the DSF@PBA-CS-PLGA NPs were confirmed using FTIR UVs, DLS, ELS, TEM, 1HNMR, DSC. Our in vitro degradation experiments showed that PBA-CS-PLGA NPs significantly improved the stability of DSF in physiological environments. Cell experiments showed that PBA-CS-PLGA NPs improved drug uptake and strongly inhibited HepG2 cell migration. A mouse tumor model was established using Dutch H22 cells. DSF@PBA-CS-PLGA NPs showed better tumor-targeting ability than DSF@PLGA NPs, with a tumor inhibition rate of more than 60%, and they induced apoptosis and inhibited neovascularization in mouse tumor tissues. Both the in vitro and in vivo experiments indicated that the DSF@PBA-CS-PLGA NPs overcame the limitations of DSF, improving the dissolution rate and stability of the drug, ultimately offering low toxicity, sustained release, and targeted delivery. These findings demonstrated the potential of DSF@PBA-CS-PLGA NPs for hepatic carcinoma therapy.

Ethylenediamine-modified alginate - A hemocompatible platform for polymer-drug conjugates

The study is dedicated to the synthesis, rheological properties, hemocompatibility, and further modification of water-soluble derivatives of sodium alginate containing fragments of ethylenediamine (Alg-EDA). Alg-EDA with an equal ratio of amide/amine groups and varying degrees of substitution were synthesized by the carbodiimide method. The influence of the molecular weight of Alg-EDA on the attachment of bioactive molecules such as hydroxybenzoic and ferulic acids was determined. Modification of alginate with ethylenediamine fragments leads to a reduction in dynamic viscosity and sensitivity to Ca2+ ions (internal gelation method). Alg-EDA derivatives, with differ in molecular weight and degree of substitution with phenolic acids, are characterized by high hemocompatibility (in vitro tests: erythrocyte hemolysis, blood recalcification time, activated partial thromboplastin time). Antioxidant properties of the synthesized alginate derivatives were characterized using models of varying complexity (including cellular models). It was found that Alg-EDA, at a concentration of 0.5 mg/mL, had a statistically significant membrane-protective activity under conditions of acute oxidative stress induced by both H2O2 and AAPH. Derivatives with lower molecular weight were more effective than high molecular weight ones. Modification of polysaccharides by the fragments of phenolic acids into the structure contributed to the enhancement of antioxidant properties. The hemocompatible macromolecular antioxidants synthesized in this study are promising for further in-depth investigation for the creation of biomedical materials.

ROS-Responsive Nanoprobes for Bimodal Imaging-Guided Cancer Targeted Combinatorial Therapy.

Chemotherapy mediated by Reactive oxygen species (ROS)-responsive drug delivery systems can potentially mitigate the toxic side effects of chemotherapeutic drugs and significantly enhance their therapeutic efficacy. However, achieving precise targeted drug delivery and real-time control of ROS-responsive drug release at tumor sites remains a formidable challenge. Therefore, this study aimed to describe a ROS-responsive drug delivery system with specific tumor targeting capabilities for mitigating chemotherapy-induced toxicity while enhancing therapeutic efficacy under guidance of Fluorescence (FL) and Magnetic resonance (MR) bimodal imaging. Indocyanine green (ICG), Doxorubicin (DOX) prodrug pB-DOX and Superparamagnetic iron oxide (SPIO, Fe3O4) were encapsulated in poly(lactic-co-glycolic acid) (PLGA) by double emulsification method to prepare ICG/ pB-DOX/ Fe3O4/ PLGA nanoparticles (IBFP NPs). The surface of IBFP NPs was functionalized with mammaglobin antibodies (mAbs) by carbodiimide method to construct the breast cancer-targeting mAbs/ IBFP NPs (MIBFP NPs). Thereafter, FL and MR bimodal imaging ability of MIBFP NPs was evaluated in vitro and in vivo. Finally, the combined photodynamic therapy (PDT) and chemotherapy efficacy evaluation based on MIBFP NPs was studied. The multifunctional MIBFP NPs exhibited significant targeting efficacy for breast cancer. FL and MR bimodal imaging clearly displayed the distribution of the targeting MIBFP NPs in vivo. Upon near-infrared laser irradiation, the MIBFP NPs loaded with ICG effectively generated ROS for PDT, enabling precise tumor ablation. Simultaneously, it triggered activation of the pB-DOX by cleaving its sensitive moiety, thereby restoring DOX activity and achieving ROS-responsive targeted chemotherapy. Furthermore, the MIBFP NPs combined PDT and chemotherapy to enhance the efficiency of tumor ablation under guidance of bimodal imaging. MIBFP NPs constitute a novel dual-modality imaging-guided drug delivery system for targeted breast cancer therapy and offer precise and controlled combined treatment options.

L-Ornithine derivatives with structural hetaryl and alkyl moiety: Synthesis and antibacterial activity

Objectives. Cationic amphiphiles and antimicrobial peptidomimetics are widely investigated as antibacterial agents due to their membrane-active mechanism of action. Particular attention is focused on the rational design of compounds in this class to achieve high antimicrobial activity. The aim of the present work is to synthesize bivalent cationic amphiphiles with L-ornithine as a branching element and evaluate the effectiveness of their antibacterial action. The compounds differ in terms of hydrophobicity due to the variation of N-terminal aliphatic amino acids in the polar block and alternation of dialkyl and alkyl-hetaryl radicals in the lipophilic block.Methods. For the synthesis of nonpolar fragments of amphiphiles, methods for the alkylation of amines with alkyl bromides in the presence of carbonate salts were used. The formation of amide bonds of L-ornithine derivatives with amino acids was carried out using the carbodiimide method. For the reaction products recovery from the reaction mixture, column chromatography on silica gel and aluminum oxide activated Brockmann Grade II was used. The antimicrobial activity of the synthesized compounds against gram-positive B. subtilis 534 and gram-negative E. coli M17 bacterial strains was evaluated. Minimum inhibitory concentration (MIC) values were recorded using a serial microdilution method in a nutrient medium.Results. Developed schemes for the preparation of bivalent cationic amphiphiles based on L-ornithine derivatives are presented. Differences in the structure of aliphatic amino acids (glycine, β-alanine, γ-aminobutyric acid (GABA)), in the length of alkyl radicals (C8, C12), or in the presence of an indole moiety, were used in the design of target compounds. The high antibacterial activity of the synthesized compounds was demonstrated. The most active compounds were lipoamino acids with terminal GABA residues and asymmetrical non-polar block (tryptamyl–dodecylamine). The MIC values were 0.39 μg/mL for gram-positive bacteria and 1.56 μg/mL for gram-negative bacteria. A GABA derivative with a symmetrical lipophilic moiety based on dioctylamine demonstrated activity with an MIC of 0.78 μg/mL against B. subtilis and 3.12 μg/mL against E. coli.Conclusions. Nine new lipoamino acid cationic bivalent amphiphiles based on L-ornithine were synthesized. The structure of the obtained compounds was confirmed by nuclear magnetic resonance 1H spectroscopy and mass spectrometry data. Leading compounds in antimicrobial activity against both gram-positive and gram-negative strains of bacteria were determined. The influence of the degree of lipophilicity in the asymmetric nonpolar block on the level of exhibited antimicrobial activity is demonstrated.

Hybrid poly(dopamine/phenosafranin) microparticles as replicas of yeast capsules for the immobilization of myoglobin

Yeast capsules (YCs) were loaded with a photosensitive phenosafranin (PSF) dye through electrostatic force-driven spontaneous deposition and served as templates for synthesizing biocompatible microparticles. Oppositely charged dopamine and PSF were encapsulated and deposited onto the yeast polysaccharide scaffold through ionic complexation. Subsequently, self-oxidative polymerization was carried out in situ, forming YC@(PDPSF) microparticles. To extend an innovative idea for diverse design and multipurpose YC@(PDPSF) applications, we immobilized myoglobin (Mb) on their surface. The multifunctional coating facilitated the immobilization of Mb involved in oxygen (O2) storage utilizing the carbodiimide method with 6-aminocaproic acid (ACA) as a linker. The biological activity of Mb encapsulated in YCs and immobilized on the surface of YC@(PDPSF) microparticles was compared using UV–vis spectrophotometry. The assays showed that the catalytic activity of Mb was sufficiently retained after immobilization. This new YC@(PDPSF)-Mb microparticles will have potential applications in biosensors, particularly for detecting and separating specific analytes and small molecules (e.g. NO, CO). Their unique properties could make them valuable for various biomedical purposes including enzyme assays, chromatography, air pollution control devices, and filtration processes in gas mask filters.

Identification of the Vulnerability of Atherosclerotic Plaques by a Photoacoustic/Ultrasonic Dual-Modal cRGD Nanomolecular Probe.

To explore the feasibility of using cRGD-GNR-PFP-NPs to assess plaque vulnerability in an atherosclerotic plaque mouse model by dual-modal photoacoustic/ultrasonic imaging. A nanomolecular probe containing gold nanorods (GNRs) and perfluoropentane (PFP) coated with the cyclic Arg-Gly-Asp (cRGD) peptide were prepared by double emulsion solvent evaporation and carbodiimide methods. The morphology, particle size, potential, cRGD conjugation and absorption features of the nanomolecular probe were characterized, along with its in vitro phase transformation and photoacoustic/ultrasonic dual-modal imaging properties. In vivo fluorescence imaging was used to determine the distribution of cRGD-GNR-PFP-NPs in vivo in apolipoprotein E-deficient (ApoE-/-) atherosclerotic plaque model mice, the optimal imaging time was determined, and photoacoustic/ultrasonic dual-modal molecular imaging of integrin αvβ3 expressed in atherosclerotic plaques was performed. Pathological assessments verified the imaging results in terms of integrin αvβ3 expression and plaque vulnerability. cRGD-GNR-PFP-NPs were spherical with an appropriate particle size (average of approximately 258.03±6.75 nm), a uniform dispersion, and a potential of approximately -9.36±0.53 mV. The probe had a characteristic absorption peak at 780~790 nm, and the surface conjugation of the cRGD peptide reached 92.79%. cRGD-GNR-PFP-NPs were very stable in the non-excited state but very easily underwent phase transformation under low-intensity focused ultrasound (LIFU) and had excellent photoacoustic/ultrasonic dual-modal imaging capability. Mice fed a high-fat diet for 20 weeks had obvious hyperlipidemia with larger, more vulnerable plaques. These plaques could be specifically targeted by cRGD-GNR-PFP-NPs as determined by in vivo fluorescence imaging, and the enrichment of nanomolecular probe increased with the increasing of plaque vulnerability; the photoacoustic/ultrasound signals of the plaques in the high-fat group were stronger. The pathological assessments were in good agreement with the cRGD-GNR-PFP-NPs plaque accumulation, integrin αvβ3 expression and plaque vulnerability results. A phase variant photoacoustic/ultrasonic dual-modal cRGD nanomolecular probe was successfully prepared and can be used to identify plaque vulnerability safely and effectively.

Using anti-PD-L1 antibody conjugated gold nanoshelled poly (Lactic-co-glycolic acid) nanocapsules loaded with doxorubicin: A theranostic agent for ultrasound imaging and photothermal/chemo combination therapy of triple negative breast cancer.

Triple negative breast cancer (TNBC) has the worst prognosis of all breast cancers, and it is difficult to progress through traditional chemotherapy. Therefore, the treatment of TNBC urgently requires agents with effective diagnostic and therapeutic capabilities. In this study, we obtained programmed death-ligand 1 (PD-L1) antibody conjugated gold nanoshelled poly(lactic-co-glycolic acid) (PLGA) nanocapsules (NCs) encapsulating doxorubicin (DOX) (DOX@PLGA@Au-PD-L1 NCs). PLGA NCs encapsulating DOX were prepared by a modified single-emulsion oil-in-water (O/W) solvent evaporation method, and gold nanoshells were formed on the surface by gold seed growth method, which were coupled with PD-L1 antibodies by carbodiimide method. The fabricated DOX@PLGA@Au-PD-L1 NCs exhibited promising contrast enhancement in vitro ultrasound imaging. Furthermore, DOX encapsulated in NCs displayed good pH-responsive and photo-triggered drug release properties. After irradiating 200 μg/mL NCs solution with a laser for 10 min, the solution temperature increased by nearly 23°C, indicating that the NCs had good photothermal conversion ability. The targeting experiments confirmed that the NCs had specific target binding ability to TNBC cells overexpressing PD-L1 molecules. Cell experiments exhibited that the agent significantly reduced the survival rate of TNBC cells through photochemotherapy combination therapy. As a multifunctional diagnostic agent, DOX@PLGA@Au-PD-L1 NCs could be used for ultrasound targeted contrast imaging and photochemotherapy combination therapy of TNBC cells, providing a promising idea for early diagnosis and treatment of TNBC.

Aptamer-functionalized polydiacetylene biosensor for the detection of three foodborne pathogens.

Rapid, simple and sensitive screening of foodborne pathogens is of great significance to ensure food safety. In this study, an aptamer-functionalized polydiacetylene (Apta-PDA) biosensor was developed for the detection of E. coli O157:H7, S. typhimurium or V. parahaemolyticus. First, aptamers responding to the target bacteria were modified on the surface of magnetic beads by covalent binding to form MBs-oligonucleotide conjugates for bacterial enrichment. Then, an Apta-PDA biosensor was obtained by connecting the aptamers to the PDA nanovesicles using the carbodiimide method. Molecular recognition occurred in the presence of the target bacteria, whereby the aptamer folded into a sequence-defined unique structure, resulting in an MBs-Apta/bacteria/Apta-PDA sandwich structure. Due to the optical properties of PDA, the blue-red transition of the detection system could be observed by the naked eye and quantified by the colorimetric response percentage (CR%). Under optimized conditions, the detection limits of E. coli O157:H7, S. typhimurium and V. parahaemolyticus were 39, 60 and 60CFU/ml, respectively, with a selectivity of 100% and a reaction time of 30min. Compared with the gold standard method, the accuracy of the three target bacteria detection reached 98%, 97.5% and 97%, respectively, and the sensitivity and specificity were both greater than 90%. The entire detection process was rapid and easy to execute without any special equipment, making this technology particularly suitable for resource-poor laboratories or regions.

Efficient Hapten-Specific Biopanning Strategy Based on the Fe3O4@ENR-Functionalized Core-Shell Magnetic Nanoparticles.

The biopanning of target-specific phages is one of the most critical steps in the preparation of single-domain antibodies. In the traditional biopanning of haptens, the nonspecific binding of library phages to macromolecular proteins is one of the most challenging problems in preparing single-domain antibodies. In this research, Fe3O4@ENR-functionalized core-shell magnetic nanoparticles (FMNPs) were silylated and aminated by tetraethyl orthosilicate and (3-aminopropyl)triethoxysilane, and target enrofloxacin was coupled onto the surface by the carbodiimide method. The magnetic nanoparticles were characterized by Fourier transform infrared spectroscopy, particle size distribution, zeta potential, transmission electron microscopy observation, and indirect enzyme-linked immunosorbent assay (ELISA). A biopanning strategy based on Fe3O4@ENR FMNPs was then established to solve the problem in the traditional solid-phase biopanning process. The results showed that a considerable number of enrofloxacin (ENR)-positive phages were screened by only one round of biopanning. Finally, two ENR-specific shark-derived single-domain genes were identified and validated by monoclonal phage ELISA, gene sequencing, and biolayer interferometry technology. Our study provides a new biopanning strategy based on Fe3O4@ENR FMNPs for efficiently providing phages specific to haptens.

Immune response of S. Typhi-derived Vi polysaccharide and outer membrane protein a conjugate in mice.

Typhoid fever is a serious concern precisely in developing nations. Still investigators are exploring a better conjugate partner for Vi-polysaccharide to develop a more effective vaccine for typhoid fever. Here, we cloned and expressed S. Typhi outer membrane protein A (OmpA). The conjugation of Vi-polysaccharide with OmpA was carried out by the carbodiimide (EDAC) method employing ADH as a linker. Total Ig and IgG generated against OmpA, and Vi polysaccharide was quantified by ELISA. Vi polysaccharide alone induced very low levels of Vi polysaccharide antibody. Vi-OmpA conjugate (Vi-conjugate) elicited a robust immune response compared to Vi polysaccharide alone and showed booster response. Further, IgG was only evoked by Vi-OmpA conjugate, not with Vi polysaccharide alone. OmpA antibody induction in both the Vi-OmpA conjugate and OmpA were similar level. Taken together, we show that OmpA as a carrier protein conjugated to Vi polysaccharide is immunogenic. We predict OmpA antibodies will contribute protection along with antibodies generated by Vi-polysaccharide. Past and current literature supports that OmpA is highly conserved protein not only among Salmonellae but entire Enterobacteriacea family with 96-100% identity.

Development of ic-ELISA and colloidal cold-based immunochromatographic assay for red 2G detection in fruit drinks, red wine, and yoghurts

Red 2G (R2G), a cheap industrial colorant, cannot be added to food. An anti-R2G monoclonal antibody (mAb) was prepared by immunizing mice with the conjugate of R2G hapten and protein, which based on the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) method. Indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and colloidal gold-based immunochromatographic assay (CG-ICA) methods were used to determine R2G in fruit drinks, red wine, and yoghurts. A standard curve of the developed ic-ELISA showed that the IC50 of the anti-R2G mAb was 1.02 ng/mL, and limit of detection value (LOD) was 0.21 ng/mL. For the CG-ICA developed, the visual limit of detection values (vLOD) were 2 ng/mL and cut-off values of 100 ng/mL in samples. The results indicated that these two methods could be used to quickly detect R2G in fruit drinks, red wine, and yoghurts.

The specific biopanning of single-domain antibody against haptens based on a functionalized cryogel.

Phage display technology is commonly applied for high-throughput screening of single-domain antibodies (sdAbs), and the problem of non-specific adsorption caused by carrier proteins seriously affects the biopanning of single-domain antibodies specific to haptens. In this paper, enrofloxacin (ENR)-functionalized cryogels were prepared by the ethylenediamine (EDA) and carbodiimide methods for application in the biopanning of ENR-specific phages. To improve the efficiency of biopanning, double blocking, a wash solution flow rate of 1 mL/min, and phage pre-incubation were applied to the biopanning process through single-factor experiments. Results of flat colony counting showed that the phage output of AG-ENR cryogels was 15 times higher than that of AG cryogels for the same input amount. And seven complete sequences of ENR-specific shark sdAbs were obtained by monoclonal phage ELISA and sequence alignment. All these results indicate that functionalized cryogels could be used as a novel and efficient method for phage biopanning for single-domain antibodies to haptens.

Antimicrobial and anti-biofilm activities of chlorogenic acid grafted chitosan against Staphylococcus aureus

In this work, Chitosan-grafted-chlorogenic acid (CS-g-CA) was prepared by the carbodiimide method. The purpose of this study was to investigate the antibacterial and anti-biofilm activity of CS-g-CA against Staphylococcus aureus (S. aureus). The minimum inhibitory concentration (MIC) of CS-g-CA against S. aureus was identified as 0.625 mg/mL. S. aureus treated with 1/2 × MIC of CS-g-CA had a longer logarithmic growth phase than that of the CK group, while 1 × MIC and 2 × MIC inhibited the growth of bacteria. The damaging effect of CS-g-CA on bacterial cells was analyzed by measuring the activity of cellular antioxidant enzymes (Catalase (CAT) and Glutathione peroxidase (GSH-Px)) and intracellular enzymes (alkaline phosphatase (AKPase) and adenosine triphosphatase (ATPase)). The results of DNA gel electrophoresis illustrated that CS-g-CA disrupted the normal metabolism of bacteria. Scanning electron microscopy (SEM) results showed that S. aureus shrank and died under CS-g-CA treatment. 1 × MIC of CS-g-CA can significantly inhibit the formation of biofilms, and 1/2 × MIC of CS-g-CA control the swimming speed of S. aureus. In addition, 77.53% mature biofilm and 60.62% extracellular polysaccharide (EPS) in the mature biofilm of S. aureus were eradicated by CS-g-CA at 2 × MIC. Confocal laser scanning microscopy (CLSM) observation further confirmed these results. Therefore, CS-g-CA was an antimicrobial and antibiofilm agent to control S. aureus, which can effectively controlling the growth of S. aureus in food, thereby preventing the occurrence of food-borne diseases.

A novel targeted iron oxide nanocarrier for inhibiting M2-type macrophages in the tumor microenvironment.

Tumor-associated macrophages (TAMs) are vital to the tumor microenvironment. They are classified as antitumor M1-type or protumor M2-type macrophages. M2-type macrophages accumulate in the tumor stroma and are related to poor prognosis. Iron oxide nanoparticles are used as drug delivery vehicles because of the structure of carboxyl groups on their surface and their ability to be easily phagocytosed by macrophages. The signal transducer and activator of transcription 6 (STAT6) signaling pathway controls M2 macrophage polarization, but the STAT6 signaling pathway inhibitor AS1517499 lacks efficient targeting in vivo. Thus, our study aimed to block the polarization of TAMs to M2-type macrophages. We used ultrasmall superparamagnetic iron oxide nanoparticles (USPIONs) as drug carriers coated with the STAT6 signaling pathway inhibitors AS1517499 and CD163 monoclonal antibodies to synthesize the targeted nanocomplex AS1517499-USPION-CD163 utilizing the carbodiimide method. Then, we determined its physicochemical properties, including hydrodynamic size distribution, ultrastructure, iron concentration, protein content and activity of the CD163 monoclonal antibody, AS1517499 content, and selectivity for M2-type macrophages, and its biological applications. The hydrodynamic size distribution was stable (average size = 95.37 nm). Regarding biological applications, the targeted nanocomplex selectively inhibited M2-type macrophages. The targeted nanocomplex AS1517499-USPION-CD163 showed high selectivity for M2-type macrophages. Therefore, iron oxide nanoparticles targeting TAMs may be an effective approach to TAM therapy.

Conjugation of Ctx(Ile21)-Ha Antimicrobial Peptides to Chitosan Ultrathin Films by N-Acetylcysteine Improves Peptide Physicochemical Properties and Enhances Biological Activity

The importance of obtaining new compounds with improved antimicrobial activity is a current trend and challenge. Some polymers such as chitosan have shown promising bactericidal properties when they are structurally modified, which is due to the binding versatility provided by their free amines. Likewise, antimicrobial peptides (AMPs) have received attention in recent years because of their bactericidal activity that is similar to or even better than that of conventional drugs, and they exhibit a low induction rate of antimicrobial resistance. Herein, the modified AMP Ctx(Ile21)-Ha-Ahx-Cys was conjugated to chitosan using N-acetylcysteine as an intermediate by the carbodiimide method. Films were prepared using protonated chitosan in 1% acetic acid and Ctx(Ile21)-Ha-Ahx-Cys AMP dissolved in N-acetylcysteine-chitosan; 1.6 mmol of ethylcarbodiimide hydrochloride, 1.2 mmol of N-hydroxysulfosucchimide, and 0.1 mol L –1of N-morpholino)ethanesulfonic acid buffer at pH 6.5 by continuous stirring at 100 × g for 10 min at 37 °C. Physicochemical properties were evaluated by Fourier-transform infrared spectroscopy, differential scanning calorimetry/thermogravimetric analysis, and X-ray diffraction to determine the mechanical properties, solubility, morphology, and thickness. Furthermore, the antimicrobial activities of chitosan-based conjugated films were evaluated againstStaphylococcus aureus,Pseudomonas aeruginosa,SalmonellaTyphimurium, andEscherichia coli. The results showed that the conjugation of a potent AMP could further increase its antibacterial activity and maintain its stable physicochemical properties. Therefore, the developed peptide–chitosan conjugate could be applied as an additive in surgical procedures to prevent and combat bacterial infection.

Comparative study of magnetic beads and microplates as supports in heterogeneous amplified assay of miRNA-141 by using mismatched catalytic hairpin assembly reaction

Magnetic beads (MBs) are often considered as an effective carrier in heterogeneous assays due to the simplicity of separation and washing, and the ability to increase and control the surface area. However, the effect of the MBs surface on the analytical parameters is poorly characterized and is often postulated from intuitive considerations. Herein, experimental evaluation through the comparison of MBs and microwell plate was carried out using the miRNA-141 (biomarker for cancer) as a target, the detection of which was performed by chemiluminescent assay with a homogeneous mismatched catalytic hairpin assembly (mCHA) reaction. The mCHA reaction produced double-stranded (ds) DNA labeled at one end with fluorescein (Flu) for capture with anti-Flu antibodies immobilized on a solid carrier, on the other end with biotin for recognition by streptavidin-polyperoxidase conjugate. The conditions of immobilization of anti-Flu antibody on MBs (a diameter of 440 nm) performed using a carbodiimide method were optimized by varying the antibody concentration in the reaction solution. It was shown that the dependence of chemiluminescent signal as a function of the concentration of anti-FluAb-MBs conjugates had a bell-shaped character. The maximum chemiluminescence was produced at the concentration of the conjugates of 2 × 109 particles/mL, with a surface area of 65 mm2. The identical surface area was used upon the assay performance with polystyrene microplates. Comparison of MBs- and microplate-assays for miRNA-141 determination showed that the obtained calibration curves and their detection limit values were the same and did not depend on the used carrier. The results showed that the choice of a carrier for heterogeneous assays should be guided by the convenience of the assay performance, not its surface area.

Gene delivery of chitosan-graft-polyethyleneimine vectors loaded on scaffolds for nerve regeneration

As an important transcription factor, c-Jun could upregulate growth factors expression in Schwann cells (SCs). Arginine-Glycine-Aspartate (RGD)-functionalized chitosan-graft-polyethyleneimine (RCP) gene vectors were prepared through the maleic anhydride & the carbodiimide methods, and electrostatically bound with c-Jun plasmids (pJUN), finally loaded on poly-L-lactic acid/silk fibroin parallel fiber films to fabricate nerve scaffold (RCP/pJUN-PSPF@PGA), which could locally deliver c-Jun plasmids into SCs via the mediation of RGD peptides, and upregulate the expression of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) in SCs. After the scaffold was bridged in sciatic nerve defect, the delivery of c-Jun plasmids from RCP/pJUN-PSPF@PGA facilitated SCs to sustain the expressions of NGF, BDNF and vascular endothelial growth factor in the injury field, promoting myelination, axonal growth and microvascular generation and nerve regeneration, muscle reinnervation and functional recovery. These results suggested that RCP/pDNA-PSPF@PGA, as an effective gene delivery platform, could provide a local gene therapy to improve nerve regeneration.

Graphene-Coated Iron Nitride Streptavidin Magnetic Beads: Preparation and Application in SARS-CoV-2 Enrichment

In this study, we prepared a streptavidin magnetic bead based on graphene-coated iron nitride magnetic beads (G@FeN-MB) and tried to use it for the enrichment of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The outer shell of our magnetic bead was wrapped with multiple graphene sheets, and there is no report on the application of graphene to the magnetic-bead-coating material. First, the graphene shell of G@FeN-MB was oxidized by a modified Hummer method so as to generate the carboxyl groups required for the coupling of streptavidin (SA) on the surface of the magnetic beads. X-ray photoelectron spectroscopy (XPS), Raman spectroscopy, Fourier transform infrared spectroscopy (FTIR), and transmission electron microscopy (TEM) were used to characterize the oxidized G@FeN-MB (GO@FeN-MB). Streptavidin was then linked to the surface of the GO@FeN-MB by coupling the amino of the streptavidin with the carboxyl on the magnetic beads by carbodiimide method; thus, the streptavidin magnetic beads (SAMBs) were successfully prepared. To prove the practicality of the SAMBs, biotinylated SARS-CoV-2 S1 antibody was linked with it to respectively capture SARS-CoV-2 Spike-protein-coupled polystyrene beads (S-PS) and pseudovirus with S-protein expressed. Microplate reader and fluorescence microscope results show that the SAMBs can effectively enrich viruses. In conclusion, the preparation of SAMBs with G@FeN-MB is feasible and has potential for application in the field of virus enrichment.

PLGA-Au-PFH-NPs-based Ultrasound Imaging in Detecting Fetal Spinal Deformities.

The study focused on the performance of ultrasound imaging in detecting fetal spinal deformities. First, the double emulsification method and the carbodiimide method were used to prepare the target Au-loaded nanorod phase-change nano-level contrast agent-PLGA-Au-PFH-NPs. After being characterized for physical and chemical properties, it was used in ultrasound imaging diagnosis. The results showed that the prepared PLGA-Au-PFH-NPs solution was a milky white suspension, the particle size detected by the laser particle sizer was (376.17±20.74) nm, and the Zeta potential was (-4.82±2.88) mV. Under the light microscope, it showed a spherical shape, uniform size distribution, and a very smooth surface. The encapsulation rate measured by the UV spectrophotometer was (80.63±4.82) %, and there was no significant difference in cell survival rate between different concentrations (P>0.05). Prenatal ultrasound in the observation group accurately diagnosed 10 cases with spinal deformities, and the diagnostic accuracy rate was 50%, including 5 cases of meningocele, 3 cases of invisible spina bifida, 1 case of myelomeningocele, and 1 case of hemivertebrae. In the control group, 7 cases were diagnosed correctly by conventional ultrasound, and the diagnosis accuracy rate was 35%, including 3 cases of meningocele, 3 cases of invisible spina bifida, and 1 case of hemivertebra. The diagnostic accuracy of the observation group was higher than that of the control group, and the difference was statistically significant (P<0.05). In conclusion, the prepared PLGA-Au-PFH-NPs had good physical and chemical properties. Ultrasound imaging based on the PLGA-Au-PFH-NPs had high accuracy in diagnosing fetal spinal deformities. To a certain extent, it provides a basis for clinical diagnosis of fetal spinal abnormality and some new ideas for ultrasound imaging diagnosis.

(Meth)acrylic esters of betulin and their copolymers with hydrophilic methacrylic esters and amides

Using the carbodiimide method for the synthesis of esters, acrylates and methacrylates of betulin were obtained, with a monoester yield of 55.0% and 70.5%, respectively. On the basis of synthesized (meth) acrylic derivatives of betulin, homopolymers and copolymers with hydrophilic monomers: 2-hydroxypropylmethacrylamide, N- (3-dimethylaminopropyl) methacrylamide, and methoxyoligo (ethylene glycol) methacrylate were obtained. The solubility of the obtained polymers in water and a number of organic solvents was studied.