Electrochemical immunosensor for ultrasensitive detection of human papillomaviruse type 16 L1 protein based on Ag@AuNPs-GO/SPA.

Human papillomaviruse type 16 (HPV16) is a high-risk serotype. As the main protective antigen protein, L1 protein is also the target protein for diagnosis. A simple label free electrochemical immunosensor (ECIS) was fabricated for ultrasensitive detection of HPV16 L1 protein in this work. Quasi-spherical Ag@Au core-shell nanoparticles on graphene oxide (Ag@AuNPs-GO) was developed as current response amplifier and characterized by UV-Vis Spectroscopy, Transmission Electron Microscopy and energy dispersive X-ray spectroscopy. Staphylococcal protein A was decorated on the modified electrode and utilized to immobilized the Fc portion of the monoclonal antibody specific for HPV16 L1 protein. Cyclic Voltammetry, Differential Pulse Voltammetry and Electrochemical Impedance Spectroscopy were used to verify the electrochemical performance and interfacial kinetic property. The increased concentration of HPV16 L1 protein led to slow electron transport and linearly decreased differential pulse voltammetry peak current with a detection limit of 0.002 ng mL-1 and a wide linear relationship in the range of 0.005-400 ng mL-1at a regression coefficient (R2) of 0.9948. Furthermore, this ECIS demonstrated acceptable accuracy with good reproducibility, stability and selectivity, suggesting a promising immunological strategy for HPV typing and early screening.

Electrochemical immunosensor nanoarchitectonics with the Ag-rGO nanocomposites for the detection of receptor-binding domain of SARS-CoV-2 spike protein.

As the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a grave threat to human life and health, it is essential to develop an efficient and sensitive detection method to identify infected individuals. This study described an electrode platform immunosensor to detect SARS-CoV-2-specific spike receptor-binding domain (RBD) protein based on a bare gold electrode modified with Ag-rGO nanocomposites and the biotin-streptavidin interaction system. The Ag-rGO nanocomposites was obtained by chemical synthesis and characterized by electrochemistry and scanning electron microscope (SEM). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to record the electrochemical signals in the electrode modification. The differential pulse voltammetry (DPV) results showed that the limit of detection (LOD) of the immunosensor was 7.2 fg mL-1 and the linear dynamic detection range was 0.015 ~ 158.5 pg mL-1. Furthermore, this sensitive immunosensor accurately detected RBD in artificial saliva with favorable stability, specificity, and reproducibility, indicating that it has the potential to be used as a practical method for the detection of SARS-CoV-2.

AP2/EREBP Pathway Plays an Important Role in Chaling Wild Rice Tolerance to Cold Stress.

Cold stress is the main factor limiting rice production and distribution. Chaling wild rice can survive in cold winters. AP2/EREBP is a known transcription factor family associated with abiotic stress. We identified the members of the AP2/EREBP transcription factor family in rice, maize, and Arabidopsis, and conducted collinearity analysis and gene family analysis. We used Affymetrix array technology to analyze the expression of AP2/EREBP family genes in Chaling wild rice and cultivated rice cultivar Pei'ai64S, which is sensitive to cold. According to the GeneChip results, the expression levels of AP2/EREBP genes in Chaling wild rice were different from those in Pei'ai64S; and the increase rate of 36 AP2/EREBP genes in Chaling wild rice was higher than that in Pei'ai64S. Meanwhile, the MYC elements in cultivated rice and Chaling wild rice for the Os01g49830, Os03g08470, and Os03g64260 genes had different promoter sequences, resulting in the high expression of these genes in Chaling wild rice under low-temperature conditions. Furthermore, we analyzed the upstream and downstream genes of the AP2/EREBP transcription factor family and studied the conservation of these genes. We found that the upstream transcription factors were more conserved, indicating that these upstream transcription factors may be more important in regulating cold stress. Meanwhile, we found the expression of AP2/EREBP pathway genes was significantly increased in recombinant inbred lines from Nipponbare crossing with Chaling wild rice, These results suggest that the AP2/EREBP signaling pathway plays an important role in Chaling wild rice tolerance to cold stress.

Design, Synthesis and Biological Evaluation of Novel PEG-Rakicidin B1 Hybrid as Clostridium difficile (CD) Targeted Anti-Bacterial Agent.

Rakicidin B1 was isolated and purified from the culture broth of a marine Streptomyces sp. as a potent anti-cancer agent, and lately the compound and its derivatives have firstly been found to possess anti-Clostridium difficile (CD) activity but with high cytotoxicity. Herein, following our previous discovery on anti-CD activity of Rakicidin B1, structure modification was performed at the OH position of Rakicidin B1 and a new Rakicidin B1-PEG hybrids FIMP2 was facilely designed and synthesized by conjugating the PEG2000 with the scaffolds of Rakicidin B1 via the linkage of carbamate. The cytotoxicity of the FIMP2 was first evaluated against three different cancer cell lines, including HCT-8 cells, PANC-1, and Caco-2, with IC50 values at 0.519 µM, 0.815 µM, and 0.586 µM, respectively. Obviously, as compared with a positive control group treated with Rakicidin B1, the IC50 value of FIMP2 increased by nearly 91-fold, 50-fold, and 67-fold, suggesting that the PEGylation strategy significantly reduced the cytotoxicity of FIMP2. Thus, this preliminary result may be beneficial to increase its safety index (SI) value due to the decreased cytotoxicity of FIMP2. In addition, this decreased cytotoxicity of FIMP2 was further confirmed based on a zebrafish screening model in vivo. Thereafter, the anti-CD activity of FIMP2 was evaluated in vivo, and its efficacy to treat CDI was found to be better than that of vancomycin. The mortality and recurrence rate of FIMP2 is not as low compared with that of vancomycin; these results demonstrated that compound FIMP2 is a new, promising anti-CD agent with significant efficacy against CD recurrence with low cytotoxicity towards bodies.

Facile Transformation from Rofecoxib to a New Near-Infrared Lipid Droplet Fluorescent Probe and Its Investigations on AIE Property, Solvatochromism and Mechanochromism.

Lipid-related cancers cause a large number of deaths worldwide. Therefore, development of highly efficient Lipid droplets (LDs) fluorescent imaging probes will be beneficial to our understanding of lipid-related cancers by allowing us to track the metabolic process of LDs. In this work, a LDs-specific NIR (λmax = 698 nm) probe, namely BY1, was rationally designed and synthesized via a one-step reaction by integrating triphenylamine (electron-donor group) unit into the structure of rofecoxib. This integration strategy enabled the target BY1 to form a strong Donor-Acceptor (D-A) system and endowed BY1 with obvious aggregation-induced emission (AIE) effect. Meanwhile, BY1 also showed observable solvent effect and reversible mechanochromatic luminescent property, which could be interpreted clearly via density functional theory (DFT) calculations, differential scanning calorimetry (DSC), powder X-ray diffraction (XPRD), and single crystal X-ray data analysis. More importantly, BY1 exhibited highly specific fluorescent imaging ability (Pearson's correlation = 0.97) towards lipid droplets in living HeLa cells with low cytotoxicity. These results demonstrated that BY1 is a new promising fluorescent probe for lipid droplets imaging, and it might be beneficial to facilitate biological research of lipid-related cancers.

Screening and identification of B cell epitope of the nucleocapsid protein in SARS-CoV-2 using the monoclonal antibodies.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus that causes the coronavirus disease (COVID-19). It is confirmed that nucleocapsid (N) protein is closely related to viral pathogenesis, modulation of host immune response, RNA transcription, and replication and virus packaging. Therefore, the N protein is a preponderant antigen target for virus detection. The codon-optimized N gene was designed according to the encoding characteristics of insect cells and inserted into pFastBacTM1 vector with 6 × His-tag-fused N protein for expression in insect sf21 cells. Six anti-N mAbs (4G3, 5B3, 12B6, 18C7-A2, 21H10-A3, 21H10-E9) were prepared by recombinant N protein. The mAbs showed high titers, antibody affinity, and reactivity with the SARS-CoV-2 N protein. Then, fourteen overlapped peptides that covered the intact N protein were synthesized (N1-N14). Peptide N14 was identified as the main linear B-cell epitope region via peptide-ELISA and dot-blot assay, and this region was truncated gradually until mapping the peptide 401-DFSKQLQQ-408. Simultaneously, compared with the sequence of variants of concern (VOCs) and variants of interest (VOIs) strains among the several countries, epitope 401-DFSKQLQQ-408 is very conservative among them. The findings provide new guidance for the design and detection of COVID-19 targets. KEY POINTS: • The N protein was optimized according to the insect cell codon preference and was highly expressed. • The monoclonal antibodies prepared in this study were shown high antibody titers and high affinity. • Monoclonal antibodies were used to map the epitope 401-408 amino acids of N protein for the first time in this study.

Differential diagnosis of the infection caused by wild-type or CD2v-deleted ASFV strains by quantum dots-based immunochromatographic assay.

African swine fever (ASF), a highly contagious and lethal disease, poses a tremendous threat and burden to the swine industry worldwide. Lack of available vaccines or treatments leaves rapid diagnosis as the key tool to control the disease. Quantum dots (QDs) are unique fluorescent semiconductor nanoparticles, highly versatile for biological applications. In this study, we developed a quantum dots-based fluorescent immunochromatographic assay (QDs-FICA) using CD2v as the diagnosis antigen to detect ASFV antibodies. The titre of the test strip was 1 : 5·12 × 105 . In addition, the strip was highly specific to anti-ASFV serum and had no cross-reaction with CSFV, PPV, PRRSV, PCV-2, PRV and FMDV. Moreover, a comparative test of 71 clinical samples showed that the coincidence rate was 85·92% between the test strip and the commercial ELISA kit (coated with p30, p62 and p72). The QDs-FICA can be used to detect ASFV antibodies, which is meaningful for the surveillance, control and purification of ASF.

Quantum dot-based fluorescence immunosorbent assay for the rapid detection of bacitracin zinc in feed samples.

Bacitracin zinc (BAC), a polypeptide antibiotic, is utilized as a feed additive due to its ability to promote growth in animals. However, the abuse of BAC can lead to a great threat to food safety. Therefore, there is an urgent need to develop a rapid and sensitive detection method. In this study, a monoclonal antibody (mAb) against BAC with excellent sensitivity and specificity was obtained. For the first time, quantum dots (QDs) were conjugated with the prepared mAb against BAC and rabbit anti-mouse antibody to fabricate a direct and an indirect competitive fluorescence-linked immunosorbent assay (dc-FLISA and ic-FLISA) to detect BAC. The IC50 of dc-FLISA and ic-FLISA were 0.28 ng/ml and 0.17 ng/ml, respectively. The limits of detection were 0.0016 ng/ml and 0.001 ng/ml, respectively, and the detection ranges were 0.0016-46.50 ng/ml and 0.001-35.65 ng/ml, respectively. In addition, the recovery rate of the two methods ranged from 93.5% to 112.0%, and the coefficient of variation (CV) was less than 10%. Therefore, the methods developed in this work have the merits of low cost, simple operation, and high sensitivity, which provide an effective analytical tool for BAC residue detection in feed samples.

A Universal Fluorescent Immunochromatography Assay Based on Quantum Dot Nanoparticles for the Rapid Detection of Specific Antibodies against SARS-CoV-2 Nucleocapsid Protein.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the pathogenic agent leading to COVID-19. Due to high speed of transmission and mutation rates, universal diagnosis and appropriate prevention are still urgently needed. The nucleocapsid protein of SARS-CoV-2 is considered more conserved than spike proteins and is abundant during the virus' life cycle, making it suitable for diagnostic applications. Here, we designed and developed a fluorescent immunochromatography assay (FICA) for the rapid detection of SARS-CoV-2-specific antibodies using ZnCdSe/ZnS QDs-conjugated nucleocapsid (N) proteins as probes. The nucleocapsid protein was expressed in E.coli and purified via Ni-NTA affinity chromatography with considerable concentration (0.762 mg/mL) and a purity of more than 90%, which could bind to specific antibodies and the complex could be captured by Staphylococcal protein A (SPA) with fluorescence displayed. After the optimization of coupling and detecting conditions, the limit of detection was determined to be 1:1.024 × 105 with an IgG concentration of 48.84 ng/mL with good specificity shown to antibodies against other zoonotic coronaviruses and respiratory infection-related viruses (n = 5). The universal fluorescent immunochromatography assay simplified operation processes in one step, which could be used for the point of care detection of SARS-CoV-2-specific antibodies. Moreover, it was also considered as an efficient tool for the serological screening of potential susceptible animals and for monitoring the expansion of virus host ranges.

Multiple fluorescence immunoassay for the simultaneous detection of Zearalenone and Ochratoxin A.

A sensitive and accurate multiple fluorescence immunoassay for the simultaneous quantitative detection of Zearalenone (ZEN) and Ochratoxin A (OTA) in single spot based on multicolor quantum dots (QDs) labeling was developed for the first time. Two kinds of ZnCdSe/ZnS (core/shell) QDs with maximum emission wavelengths at 520 nm (green) and 610 nm (orange-red) were selected as marking materials, respectively. The anti-ZEN-mAb-QDs and anti-OTA-mAb-QDs were designed as the immune fluorescent probes. Fluorescence was measured at the same excitation wavelength and two different emission wavelengths to determine each target. The procedure for QDs-based multiple fluorescence labeled immunosorbent assay (M-FLISA) was developed. The 50% inhibition concentrations (IC50) of ZEN and OTA were 0.034 and 1.175 ng/mL. Moreover, the limits of detection (LOD) for the simultaneous determination were 0.0239 and 2.339 ng/g for ZEN and OTA in maize, respectively. In addition, the recoveries ranged from 93.15 to 101.90% for ZEN and from 95.29 to 102.43% for OTA, with the coefficient variation (CV) of 2.70-8.86% and 3.51-6.22% respectively. There was good consistency between the M-FLISA and high performance liquid chromatography (HPLC) results, which confirmed that the M-FLISA was suitable for the simultaneous quantitative detection of various mycotoxins.

Detection of ochratoxin A by quantum dots-based fluorescent immunochromatographic assay.

Ochratoxin A (OTA) is a toxic metabolite produced mainly by Aspergillus and Penicillium species. A quantitative method was developed for the rapid, simple, and sensitive detection of OTA in corn by quantum dots-based fluorescent immunochromatographic assay (QDs-ICA). The CdSe/ZnS QDs-labelled anti-OTA monoclonal antibody (mAb) conjugates were designed as the fluorescent signal probe. The QDs-ICA included the designation of test line (T line) and control line (C line), which were sprayed with optimal concentrations of the OTA-OVA and staphylococcal protein-A (SPA), respectively. Under the optimal experimental conditions, the QDs-ICA exhibited excellent specificity and good accuracy and precision. For qualitative detection, the cut-off value for the T line of the visual detection method was 2.5 ng/mL. For quantitative detection, the linear regression equation of the standard curve was y = 0.366x + 0.514 with a reliable correlation coefficient (R2 = 0.992). Moreover, the 50% inhibition value (IC50) of the QDs-ICA was 0.91 ng/mL, the limit of detection (LOD) was 0.07 ng/mL, and the detection range was 0.05 to 10 ng/mL. In addition, the recovery rates ranged from 91.82 to 100.35% with a coefficient of variation (CV) below 3% for intra-assay, whereas the recovery rates for inter-assay changed from 94.29 to 104.62% with a CV below 10%. These results indicate that the QDs-ICA can serve as a potential large-scale preliminary device for rapid determination of OTA. Using CdSe/ZnS QDs as the fluorescent signal for quantum dots-based fluorescent immunochromatographic assay, the QDs-ICA provided a novel method for the rapid simultaneous qualitative and quantitative determination of OTA.

An Electrochemical Immunosensor Based on SPA and rGO-PEI-Ag-Nf for the Detection of Arsanilic Acid.

A sensitive electrochemical immunosensor was prepared for rapid detection of ASA based on arsanilic acid (ASA) monoclonal antibody with high affinity. In the preparation of nanomaterials, polyethyleneimine (PEI) improved the stability of the solution and acted as a reducing agent to generate reduced graphene oxide (rGO) with relatively strong conductivity, thereby promoting the transfer of electrons. The dual conductivity of rGO and silver nanoparticles (AgNPs) improved the sensitivity of the sensor. The synthesis of nanomaterials were confirmed by UV-Vis spectroscopy, X-ray diffraction, transmission electron microscopy and scanning electron microscopy. In the optimal experiment conditions, the sensor could achieve the detection range of 0.50-500 ng mL-1 and the limit of detection (LOD) of 0.38 ng mL-1 (S/N = 3). Moreover, the sensor exhibited excellent specificity and acceptable stability, suggesting that the proposed sensor possessed a good potential in ASA detection. Thus, the as-prepared biosensor may be a potential way for detecting other antibiotics in meat and animal-derived foods.

Enhanced Li Storage Stability Induced by Locating Sn in Metal-Organic Frameworks.

By locating elemental Sn in an open anionic framework, the particle cracking arising from huge volume expansion of Sn-based anode materials during lithiation/delithiation is alleviated, and the cycling stability is greatly improved. The Sn-based metal-organic framework anode material shows superior cyclic stability, with a capacity retention >92 % (after 200 cycles) and high lithium storage capacity (610 mAh g-1 ).

Mitogen-activated protein kinase p38 induces HDAC4 degradation in hypertrophic chondrocytes.

Histone deacetylase 4 (HDAC4) is a critical negative regulator for chondrocyte hypertrophy by binding to and inhibiting Runx2, a critical transcription factor for chondrocyte hypertrophy. It is unclear how HDAC4 expression and stability are regulated during growth plate development. We report here that inhibition of mitogen-activated protein kinase (MAPK) p38 by dominant negative p38 or p38 inhibitor prevents HDAC4 degradation. Mutation of a potential caspase-2 and 3 cleavage site Asp289 stabilizes HDAC4 in chondrocytes. In contrast, constitutively active MAPK kinase 6 (constitutive activator of p38) transgenic mice exhibit decreased HDAC4 content in vivo. We also observed that p38 stimulates caspase-3 activity in chondrocytes. Inhibition of p38 or caspases reduced HDAC4 degradation. HDAC4 inhibited Runx2 promoter activity in a dose-dependent manner and caspase inhibitors further enhanced this inhibition by preventing HDAC4 degradation. Overall, these results demonstrate that p38 promotes HDAC4 degradation by increasing caspase-mediated cleavage, which releases Runx2 from a repressive influence of HDAC4 and promotes the chondrocyte hypertrophy and bone formation.

Indian Hedgehog, a critical modulator in osteoarthritis, could be a potential therapeutic target for attenuating cartilage degeneration disease.

The Hedgehog (Hh) family of proteins consists of Indian hedgehog (Ihh), sonic hedgehog (Shh), and desert hedgehog (Dhh). These proteins serve as essential regulators in a variety of developmental events. Ihh is mainly produced and secreted by prehypertrophic chondrocytes and regulates chondrocyte hypertrophy and endochondral bone formation during growth plate development. Tissue-specific deletion of the Ihh gene (targeted by Col2a1-Cre) causes early lethality in mice. Transgenic mice with induced Ihh expression exhibit increased chondrocyte hypertrophy and cartilage damage resembling human osteoarthritis (OA). During OA development, chondrocytes recapitulate the differentiation process that happens during the fetal status and which does not occur to an appreciable degree in adult articular cartilage. Ihh expression is up-regulated in human OA cartilage, and this upregulation correlates with OA progression and changes in chondrocyte morphology. A genetic study in mice further showed that conditional deletion of Ihh in chondrocytes attenuates OA progression, suggesting the possibility that blocking Ihh signaling can be used as a therapeutic approach to prevent or delay cartilage degeneration. However, Ihh gene deletion is currently not a therapeutic option as it is lethal in animals. RNA interference (RNAi) provides a means to knockdown Ihh without the severe side effects caused by chemical inhibitors. The currently available delivery methods for RNAi are nanoparticles and liposomes. Both have problems that need to be addressed. In the future, it will be necessary to develop a safe and effective RNAi delivery system to target Ihh signaling for preventing and treating OA.

Indian hedgehog in synovial fluid is a novel marker for early cartilage lesions in human knee joint.

To determine whether there is a correlation between the concentration of Indian hedgehog (Ihh) in synovial fluid (SF) and the severity of cartilage damage in the human knee joints, the knee cartilages from patients were classified using the Outer-bridge scoring system and graded using the Modified Mankin score. Expression of Ihh in cartilage and SF samples were analyzed with immunohistochemistry (IHC), western blot, and enzyme-linked immunosorbent assay (ELISA). Furthermore, we detected and compared Ihh protein levels in rat and mice cartilages between normal control and surgery-induced osteoarthritis (OA) group by IHC and fluorescence molecular tomography in vivo respectively. Ihh expression was increased 5.2-fold in OA cartilage, 3.1-fold in relative normal OA cartilage, and 1.71-fold in OA SF compared to normal control samples. The concentrations of Ihh in cartilage and SF samples was significantly increased in early-stage OA samples when compared to normal samples (r = 0.556; p < 0.001); however, there were no significant differences between normal samples and late-stage OA samples. Up-regulation of Ihh protein was also an early event in the surgery-induced OA models. Increased Ihh is associated with the severity of OA cartilage damage. Elevated Ihh content in human knee joint synovial fluid correlates with early cartilage lesions.

Development of quantum dot-based immunochromatographic strip for detection of antibodies against ASFV pp62.

ASFV is the only known double-stranded insect-borne DNA virus, which can rapidly infect domestic pigs and wild boars with ticks as transmission medium. Since it was first discovered in 1921, it quickly spread to all parts of the world and brought huge economic losses to the pig industry all over the world. At present, there is still no safe and effective vaccine for ASFV. Here, we developed a quantum-dot labeled antibody test strip for the detection of antibodies against ASFV pp62. The pp62 protein was labeled with quantum dots, and the antibody test strip was developed uses it in a detection mode of labeled antigen-SPA interceptor-monoclonal antibody quality control. The test strip showed high sensitivity, the positive detection limit of the strip was 1: 106 by continuous multiple dilution using the positive standard serum of ASFV antibody as reference. The test strip showed good specificity, and there was no cross reaction with other swine diseases virus (PCV2, PRRSV, CSFV, PPV). Using the detection results of commercialized kit for African swine fever virus as reference, 80 ASFV antibody negative serum and 4 different ASFV antibody positive serum were detected using the ASFV pp62 quantum-dot labeled antibody test strip. The results were consistent with the commercial kit. This study provides a new detection method for the prevention and control of African swine fever.

Fluorescence immunochromatographic detection of antibodies to the p72 protein of African swine fever virus.

The African swine fever virus (ASFV), a highly contagious pathogen responsible for African swine fever (ASF), causes significant economic losses in the global pork industry. Due to its large and complex structure, ASFV remains refractory to commercial vaccine development, necessitating the creation of rapid, sensitive, and specific diagnostic tools for disease control. In this study, quantum dots were conjugated to ASFV p72 protein to establish a fluorescent immunochromatographic assay for detecting ASFV-specific antibodies. The assay test strips contained four adjacent pads arranged sequentially: a sample-application pad, a pad containing mobile antigen-probe conjugate, a nitrocellulose readout pad featuring a test line containing immobilised staphylococcal protein A and a control line containing immobilised monoclonal antibodies against the ASFV p72 protein, and an absorbent pad driving the directional flow of liquid via capillary action. The resulting fluorescence immunochromatographic assay demonstrated highly sensitive and specific ASFV antibody detection in under 15 min. Specificity testing showed no cross-reactivity with serum antibodies against other viruses and sensitivity surpassing that of commercial ASFV antibody colloidal gold immunochromatographic test strips. This novel approach offers rapid detection, excellent specificity, and high sensitivity, and supports the future development of fluorescent immunochromatographic test strips for ASFV antibody detection.

Thiolated chitosan encapsulation constituted mucoadhesive nanovaccine confers broad protection against divergent influenza A viruses.

Influenza A virus (IAV) poses a significant threat to human and animal health, necessitating the development of universal influenza vaccines that can effectively activate mucosal immunity. Intranasal immunization has attracted significant attention due to its capacity to induce triple immune responses, including mucosal secretory IgA. However, inducing mucosal immunity through vaccination is challenging due to the self-cleansing nature of the mucosal surface. Thiolated chitosan (TCS) were explored for mucosal vaccine delivery, capitalizing on biocompatibility and bioadhesive properties of chitosan, with thiol modification enhancing mucoadhesive capability. The focus was on developing a universal nanovaccine by utilizing TCS-encapsulated virus-like particles displaying conserved B-cell and T-cell epitopes from M2e and NP proteins of IAV. The optimal conditions for nanoparticle formation were investigated by adjusting the thiol groups content of TCS and the amount of sodium tripolyphosphate. The nanovaccine induced robust immune responses and provided complete protection against IAVs from different species following intranasal immunization. The broad protective effect of nanovaccines can be attributed to the synergistic effect of antibodies and T cells. This study developed a universal intranasal nanovaccine and demonstrated the potential of TCS in the development of mucosal vaccines for respiratory infectious diseases.

Development of a double-antibody sandwich enzyme-linked immunosorbent assay for rapid detection of VZV.

BACKGROUND: Varicella zoster virus (VZV) is the pathogen of varicella and herpes zoster, it is necessary to develop a rapid, sensitive and specific detection method for the prevention and control of related diseases. METHODS: We inserted the gB protein extracellular region gene (gB-ex, 1-2208 bp) of VZV into lentivirus vector, and then obtained the recombinant gB protein through mammalian expression system. BALB/c mice were immunized multiple times with purified gB protein as immunogen. Then four strains of high affinity monoclonal antibodies targeting gB protein were prepared by cell fusion technique. Monoclonal antibodies 5G4 and HRP-4E9 were selected as capture and detection antibodies respectively, and a double-antibody sandwich ELISA method was established for detection. RESULTS: The detection limit of the DAS-ELISA was 156 PFU/mL, and there was no cross-reaction with Herpes simplex virus-1/Herpes simplex virus-2/Pseudorabies virus. The coefficients of variation of intra-assay and inter-assay repeatability were less than 5%. CONCLUSIONS: In this study, a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was established for the detection of VZV. The assay has good sensitivity, specificity and repeatability, which provides strong technical support and product guarantee for the rapid clinical detection of VZV.