Ovarian cancer remains the deadliest gynecological malignancy, largely because early screening is inadequate. This work reports a smartphone-based fluorescent homogeneous immunoassay sensor (SFHIS) that combines aggregation-induced emission luminogen-doped metal-organic framework microspheres (AMOFs) as donors and spiky carbon nanospheres (CNPs) as acceptors. Target antigen-antibody sandwich formation brings donor and acceptor into proximity, inducing efficient Förster resonance energy transfer (FRET) and fluorescence quenching, which is linearly related to the natural logarithm of antigen concentration. A dual-correction algorithm removes internal filter effects, enabling accurate quantification. The SFHIS shows linear ranges of 0.8-51.2 U/mL for CA125 and 8-256 pmol/L for HE4, with limits of detection of 0.67 U/mL and 1.03 pmol/L, respectively. The assay is completed within 25 min without washing, exhibits intra- and inter-batch CVs < 10%, no cross-reactivity with CEA, AFP, or PSA, and correlates strongly with chemiluminescence assays (CA125: R2 = 0.970; HE4: R2 = 0.982; P < 0.05). This portable, low-cost platform offers rapid, reliable biomarker detection suitable for clinical laboratories, primary care, community health, and resource-limited settings.

NK cells engineered to express interleukin-15 (IL-15) and a CD19-targeted chimeric antigen receptor (CAR) have been used to treat patients with relapsed and/or refractory B cell malignances, demonstrating encouraging outcomes and favorable safety profile. However, the effect of IL-21 in CAR-NK cell therapy remains unknown. CD19-specific CAR with 4-1BB costimulatory domain and cytokine IL-21 or IL-15 was constructed and transduced into peripheral blood (PB)-derived NK cells to produce CD19-CAR-IL21 NK cells (CAR-21) or CD19-CAR-IL15 NK cells (CAR-15), respectively. The phenotypic profile, transcriptomic characteristics, functionality and anti-tumor activity of CAR-21 NK cells and CAR-15 NK cells were compared. Compared with CAR-NK cells co-expressing IL-15, CAR-NK cells co-expressing IL-21 exhibited significantly increased IFN-γ, TNF-α and Granzyme B production, as well as degranulation, in response to CD19+ Raji lymphoma cells, resulting in enhanced cytotoxic activity upon repetitive tumor stimulation. Furthermore, IL-21 co-expression improved the in vivo persistence of CAR-NK cells and significantly suppressed tumor growth in a xenograft Raji lymphoma murine model, leading to prolonged survival of CD19+ tumor-bearing mice. RNA sequencing revealed that CAR-21 NK cells have a distinct transcriptomic signature characterized by enriched in cytokine, cytotoxicity, and metabolic related signaling, when compared with CAR-15 NK or CAR NK cells. This study demonstrated that CD19-specific CAR-NK cells engineered to express IL-21 exhibit superior persistence and anti-tumor activity against CD19+ tumor compared to CAR-NK cells co-expressing IL-15, which might be a promising therapeutic strategy for treating patients with relapse or refractory B cell malignances.

The level of rabies virus neutralizing antibody (RVNA) is a critical indicator to evaluate the protective immunity of vaccinees against rabies virus (RABV) infection. The fluorescent antibody virus neutralization test (FAVN) and the rapid fluorescent focus inhibition test (RFFIT) are the WHO recommended assays for determining the RVNA level after vaccination, but both methods are complicated and time-consuming for 1–5 days. Here we developed a smartphone-based immunosensor platform (SPIS), intelligently integrated with the nanozyme (Au@PtNPs) engaged competitive immunoassay (CIA) in the heavy ion microporous membrane (HIM) filtration microplate and PMMA optical fiber signal transmission (SPIS-CIV) for quantitatively surrogate virus neutralization testing (sVNT) of the surrogate RVNA (sRVNA) in blood samples during 50 min. The highest intra-assay and inter-assay coefficient variations (CV) of SPIS-CIA varied < 13 % or 14 %, respectively. The linear range of quantification was between 0.4 IU/mL and 10 IU/mL RVNA. The nearly full agreement was found between SPIS-CIA and FAVN for measuring sRVNA or RVNA titers in serum/plasma samples from 27 rabies vaccinees and 64 non-rabies vaccinees (R2=0.9941, P < 0.0001), especially SPIS-CIV was accurate for quantification of sRVNA at the border line of protection level (0.5 IU/mL). The limit of detection or quantification (LOD/LOQ) was determined to be 0.25 IU/mL and 0.4 IU/mL, respectively. In conclusion, SPIS-CIA is a rapid, quantitative and precise point-of-care testing for serum sRVNA level, which can be used to predict the protection efficacy against RABV infection in human rabies vaccinees in clinical diagnosis.

The level of rabies virus neutralizing antibody (RVNA) is a critical indicator to evaluate the protective immunity of vaccinees against rabies virus (RABV) infection. The fluorescent antibody virus neutralization test (FAVN) and the rapid fluorescent focus inhibition test (RFFIT) are the WHO recommended assays for determining the RVNA level after vaccination, but both methods are complicated and time-consuming. Here we developed a competitive immunoassay with Au@PtNPs nanozyme substrate development on the track-etched microporous membrane (TEM) filtration microplate and optical fibers transmitted immunosensing smartphone reader platform (TEMFIS) for quantitatively surrogate virus neutralization testing (sVNT) of RVNA in blood samples. Highest intra-assay and inter-assay coefficient variations (CV) of TEMFIS-sVNT varied < 13% or 14%, respectively. The linear range of quantification was between 0.5 IU/ml and 10 IU/ml RVNA. The nearly full agreement was found between TEMFIS-sVNT and FAVN for measuring RVNA titers in serum samples from 25 rabies vaccinees and 64 non-rabies vaccinees (R2=0.9941, P<0.0001). In conclusion, TEMFIS-sVNT is a rapid and precise point-of-care testing for quantifying serum RVNA level and predicting its protection efficacy against RABV infection in human rabies vaccinees.

Abstract Epstein-Barr virus (EBV) related post-transplant lymphoproliferative disorder (EBV-PTLD) is a life-threatening complication after hematopoietic stem cell transplantation (HSCT) or solid organ transplantation (SOT), for which no standard therapeutic means have been developed. Significant increase expression of natural killer group 2 member D ligands (NKG2DLs) was observed on B-lymphoblastoid cells of EBV-PTLD, indicating NKG2DLs as potential therapeutic targets for treatment of EBV-PTLD. In this study, the recombinant constructs of NKG2D CAR and IL-15/IL-15Rα-NKG2D CAR were generated with a retroviral vector and then transduced to human T cells to produce NKG2D CAR-T and IL-15/IL-15Rα-NKG2D CAR-T cells, respectively. B-lymphoblastoid cell lines (B-LCLs) and the xenografted mouse models were established to evaluate the efficacy of these CAR-T cells. IL-15/IL-15Rα-NKG2D CAR-T cells exhibited superior proliferation and antigen-specific cytotoxic effect compared to NKG2D CAR-T, as IL-15/IL-15Rα signaling promoted the expansion of less differentiated central memory T cells (TCM) and increased expression of CD107a and IFN-γ. Moreover, EBV DNA load was dramatically reduced, and 80% B-LCL cells were eliminated by IL-15/IL-15Rα-NKG2D CAR-T cells after co-culturing. In-vivo study confirmed that IL-15/IL-15Rα-NKG2D CAR-T cell therapy significantly enhanced antiviral efficacy in mice, as the serum load of EBV after IL-15/IL-15Rα-NKG2D CAR-T cell infusion was 1500 times lower than the untreated control (P < 0.001). The enhanced efficacy of IL-15/IL-15Rα-NKG2D CAR T cells was probably due to the IL-15/IL-15Rα signaling improved homing and persistence of NKG2D CAR-T cells in vivo, and increased the production of IFN-γ, Perforin, and Granulysin. In conclusion, NKG2D CAR-T cells co-expressing IL-15/IL-15Rα promoted the central memory CAR T cell proliferation and improved the homing and persistence of CAR T cells in vivo, resulting in enhanced anti-tumor and anti-viral effects in treating EBV-PTLD.

Toxoplasmosis is a common zoonotic disease caused by a protozoan parasite Toxoplasma gondii (Tox), approximately infecting one-third of human populations worldwide. This study developed the carbon nanospheres (CNPs) based dual spectral-overlapped fluorescence quenching lateral flow immunoassay (CNPs-FQLFIA) for detection of Tox antibodies (ToxAbs). The CNPs have been effectively coupled with Tox antigen (ToxAg), which can completely overlap the excitation and emission spectra of europium nanospheres (EuNPs) and CdSe/ZnS quantum dots (QDs) in testing strips of CNPs-QDs-FQLFIA or CNPs-EuNPs-FQLFIA. The sensitivity of CNPs-EuNPs-FQLFIA or CNPs-QDs-FQLFIA was 4 or 8 IU/mL under natural light readout, or both 4 IU/mL ToxAbs under ultraviolet (UV) light readout by the naked eyes, respectively. The limit of detection (LOD) of two types of CNPs-FQLFIA was both 1 IU/mL ToxAbs under UV light by a dry fluorescence analyzer, but no cross-reaction was found with other antibodies. The intra-assay coefficient variation (CV) of both CNPs-EuNPs-FQLFIA and CNPs-QDs-FQLFIA was less than 8%, while the inter-assay CV was less than 14%, respectively. The correlation coefficient (R2) of CNPs-EuNPs-FQLFIA or CNPs-QDs-FQLFIA to measure the different concentrations of ToxAbs spiked serum samples was 0.99712 and 0.99896, respectively. The CNPs-FQLFIA presented a characteristics of 94.3% sensitivity, 100% specificity and 98% accuracy for detection of ToxAbs in clinical serum samples. In conclusion, CNPs-FQLFIA with EuNPs or QDs fluorescence reporter was an easy, rapid, sensitive, precise and quantitative assay for detecting Tox antibodies in human blood samples.

Background: Hepatitis C virus (HCV) vaccines are an urgent need to prevent hepatitis C and its further progression of hepatocellular carcinoma. Since the promising T cell based chimpanzee adenovirus and modified vaccinia virus Ankara vectorial HCV vaccines were failed in clinical phase II trial, the vaccine designs to improve pro-tection efficacy in combination of cellular and humoral immunity have been hypothesized against multi-genotypic HCV.Methods: Eight HCV vaccine strains were constructed with two novel adenovirus vectors (Sad23L and Ad49L) encoding E1E2 or NS3-5B proteins of HCV genotype (Gt) 1b and 6a isolates, covering 80 % HCV strains prevalent in south China and south-east Asia. Eight HCV vaccine strains were grouped into Sad23L-based vaccine cocktail-1 and Ad49L-based vaccine cocktail-2 for vaccinating mice, respectively.Results: The immunogenicity of a single dose of 107-1010 PFU HCV individual vaccines was evaluated in mice, showing weak specific antibody to E1 and E2 protein but a dose-dependent T cell response to E1E2/NS3-5B peptides, which could be significantly enhanced by boosting with an alternative vector vaccine carrying ho-mologous antigen. Prime-boost vaccinations with vaccine cocktail-1 and cocktail-2 induced significantly higher cross-reactive antibody and stronger T cell responses to HCV Gt-1b/6a. The high frequency of intrasplenic and intrahepatic NS31629-1637 CD8+ T cell responses were identified, in which the high proportion of TRM and TEM cells might play an important role against HCV infection in liver.Conclusions: Prime-boost regimens with HCV vaccine cocktails elicited the broad cross-reactive antibody and robust T cell responses against multi-genotypic HCV in mice.

In one-carbon metabolism, methionine (Met) is methyl donor and folic acid (FA) is responsible for the transfer of methyl groups. Approximately 25 % of coated methionine (CMet) and coated folic acid (CFA) released in the rumen at 24 h of incubation. Therefore, the present study investigated the effects of CMet or/and CFA on growth performance, nutrient digestibility and rumen fermentation in bulls. Forty Simmental bulls, 464 +/- 15.9 kg of body weight (BW), were blocked by BW and randomly assigned to four groups in a 2 x 2 factorial arrangement of treatment, with factors being CFA 0 or 6 mg FA/kg dietary dry matter (DM) and CMet 0 or 0.66 g Met/kg dietary DM. The experiment period was 81 days long with 20 days for adaptation and 61 days for data and sample collection. Dry matter intake increased (P = 0.001) with CFA or CMet addition. The CFA x CMet interaction was noticed on average daily gain (ADG; P = 0.001) and feed conversion ratio (FCR; P = 0.020); the improvements in ADG and FCR were greater for supplementing CMet in diets with CFA than in diets without CFA. The digestibility of DM, organic matter, neutral detergent fiber and acid detergent fiber increased (P < 0.050) with CFA or CMet addition. When CMet was supplemented, crude protein digestibility increased in CFA diets and was unchanged in diets without CFA. Ruminal pH decreased (P < 0.050), but total volatile fatty acid concentration increased (P < 0.050) with CFA or CMet addition. Acetate to propionate ratio was unchanged with CFA addition, but decreased (P = 0.039) with CMet addition. Rumen ammonia-N concen-tration decreased (P < 0.050) for either CFA or CMet addition. The activities of carboxymethyl cellulase, protease, alpha-amylase and cellobiase and populations of protozoa, total bacteria, fungi, dominant cellulolytic bacteria, Ruminobacter amylophilus and Butyrivibrio fibrisolvens increased (P < 0.050) with CFA or CMet supplementation. The CFA x CMet interaction was noticed on Ruminococcus albus population (P = 0.014); when CMet was supplemented, R. albus population increased for diets without CFA and was unchanged for diets with CFA. The activities of chymotrypsin in duodenum increased (P < 0.050) with CFA or CMet supplementation, and that of amylopsin and trypsin in ileum increased (P < 0.050) for bulls receiving CFA addition. The concentrations of glucose, albumin and urea nitrogen in blood were unchanged and homocysteine decreased (P < 0.050) with CFA or CMet supplementation. The CFA x CMet interaction was noticed on blood total protein concentration (P = 0.045); when CMet was added, blood total protein concentration increased for diets without CFA and was unchanged for diets with CFA. The concentration of folate in blood increased (P = 0.001) for CFA addition. The concentration of Met increased (P = 0.002) for CMet supplementation. The results indicated that dietary CFA or/and CMet supplementation improved growth performance, nutrient digestibility and rumen fermen-tation, and the greatest daily gain was observed for combined addition of CFA and CMet in bulls.

The COVID-19 pandemic, caused by the SARS-CoV-2 virus, has had and continues to have a significant impact on global public health. One of the characteristics of SARS-CoV-2 is a surface homotrimeric spike protein, which is primarily responsible for the host immune response upon infection. Here we present the preclinical studies of a broadly protective SARS-CoV-2 subunit vaccine developed from our trimer domain platform using the Delta spike protein, from antigen design through purification, vaccine evaluation and manufacturability. The pre-fusion trimerized Delta spike protein, PF-D-Trimer, was highly expressed in Chinese hamster ovary (CHO) cells, purified by a rapid one-step anti-Trimer Domain monoclonal antibody immunoaffinity process and prepared as a vaccine formulation with an adjuvant. Immunogenicity studies have shown that this vaccine candidate induces robust immune responses in mouse, rat and Syrian hamster models. It also protects K18-hACE2 transgenic mice in a homologous viral challenge. Neutralizing antibodies induced by this vaccine show cross-reactivity against the ancestral WA1, Delta and several Omicrons, including BA.5.2. The formulated PF-D Trimer is stable for up to six months without refrigeration. The Trimer Domain platform was proven to be a key technology in the rapid production of PF-D-Trimer vaccine and may be crucial to accelerate the development and accessibility of updated versions of SARS-CoV-2 vaccines.

ABSTRACT The failure of COVID-19 vaccines to prevent SARS-CoV-2 infection and transmission, a possibly critical reason was the lack of protective mucosal immunity in the respiratory tract. Here, we evaluated the effects of mucosal and systemic immunity from a novel simian adenovirus-vectored COVID-19 vaccine (Sad23L-nCoV-S) in mice in comparison with Ad5-nCoV-S by intranasal (IN) drip and intramuscular (IM) injection vaccinations. As good as the well-known Ad5-nCoV-S vaccine, a single-dose IN inoculation of 1 × 109 PFU Sad23L-nCoV-S vaccine induced a similar level of IgG S-binding antibody (S-BAb) and neutralizing antibody (NAb) and higher IgA in serum, while IN route raised significantly higher IgG and IgA S-BAb and NAb in bronchoalveolar lavage (BAL), and specific IFN-γ secreting T-cell response in lung compared with IM route, but lower T-cell response in spleen. By prime-boost vaccination regimens with different combinations of IN and IM inoculations of Sad23L-nCoV-S vaccine, the IN-involved vaccination stimulated higher protective mucosal or local immunity in BAL and lung, while the IM-involved immunization induced higher systemic immunity in serum and spleen. A long-term sustained mucosal and systemic NAb and T- cell immunity to SARS-CoV-2 was maintained at high level over 32 weeks by prime-boost vaccination regimens with IN and IM routes. In conclusion, priming or boosting immunization with IN inoculation of Sad23L-nCoV-S vaccine could induce effective mucosal immunity and in combination of IM route could additionally achieve systemic immunity, which provided an important reference for vaccination regimens against respiratory virus infection. IMPORTANCE The essential goal of vaccination is to generate potent and long-term protection against diseases. Several factors including vaccine vector, delivery route, and boosting regimen influence the outcome of prime-boost immunization approaches. The immunization regimens by constructing a novel simian adenovirus-vectored COVID-19 vaccine and employing combination of intranasal and intramuscular inoculations could elicit mucosal neutralizing antibodies against five mutant strains in the respiratory tract and strong systemic immunity. Immune protection could last for more than 32 weeks. Vectored vaccine construction and immunization regimens have positively impacted respiratory disease prevention.

Hepatitis B surface antigen-negative, hepatitis B virus (HBV) DNA-positive occult HBV infection status appears associated with liver fibrosis; hepatitis B core antigen is correlated with elevated serum liver fibrosis indexes and may affect fibrotic progression in liver tissue. Background The impact of hepatitis B surface antigen (HBsAg)-negative/hepatitis B virus (HBV) DNA-positive occult HBV infection (OBI) on the severity of liver fibrosis remains unclear. Methods A total of 1772 patients negative for HBsAg but positive for antibody to hepatitis B core antigen (HBcAg), stratified by the presence or absence of OBI, were selected for long-term carriage leading to elevation of >= 2 of 4 liver fibrosis indexes-hyaluronic acid (HA), laminin, type III procollagen peptide (PCIII), and type IV collagen (CIV)-at testing in a Chinese hospital. Patients were tested for serum viral load, HBV markers, and histopathological changes in liver biopsy specimens. Results OBI was identified in 148 patients with liver fibrosis (8.4%), who had significantly higher levels of HA, laminin, PCIII, and CIV than 1624 fibrotic patients without OBI (P < .05). In 36 patients with OBI who underwent liver biopsy, significant correlations were observed between OBI viral load and serum HA levels (P = .01), PCIII levels (P = .01), and pathological histological activity index (HAI) scores (P < .001), respectively; HAI scores and PCIII levels (P = .04); HBcAg immunohistochemical scores and HA levels (P < .001); and HBcAg immunohistochemical scores and PCIII levels (P = .03). Positive fluorescent in situ hybridization results were significantly more frequent in patients with OBIs (80.6% vs 37.5% in those without OBIs). Among patients with OBIs, HBcAg was detected in the liver tissue in 52.8% and HBsAg in 5.6%. Conclusions OBI status appears to be associated with liver fibrosis severity.

Kidney fibrosis, characterized by the activation and expansion of the matrix-producing fibroblasts, is the common outcome of chronic kidney disease (CKD). While fibroblast proliferation is well studied in CKD, little is known about the regulation and mechanism of fibroblast depletion. Here, we show that exosomes derived from stressed/injured tubules play a pivotal role in dictating fibroblast apoptosis and fate. When human kidney tubular cells (HK-2) were stimulated with TGF-β1, they produced and released increased amounts of exosomes (TGFβ-Exo), which prevented renal interstitial fibroblasts from apoptosis. In vivo, injections of TGFβ-Exo promoted renal fibroblast survival, whereas blockade of exosome secretion accelerated fibroblast apoptosis in obstructive nephropathy. Proteomics profiling identified the tumor necrosis factor-α-induced protein 8 (TNFAIP8) as a key component enriched in TGFβ-Exo. TNFAIP8 was induced in renal tubular epithelium and enriched in the exosomes from fibrotic kidneys. Knockdown of TNFAIP8 in tubular cells abolished the ability of TGFβ-Exo to prevent fibroblast apoptosis. In vivo, gain- or loss- of TNFAIP8 prevented or aggravated renal fibroblast apoptosis after obstructive injury. Mechanistically, exosomal-TNFAIP8 promoted p53 ubiquitination leading to its degradation, thereby inhibiting fibroblasts apoptosis and inducing their proliferation. Collectively, these results indicate that tubule-derived exosomes play a critical role in controlling the size of fibroblast population during renal fibrogenesis through shuttling TNFAIP8 to block p53 signaling. Strategies to target exosomes may be effective strategies for the therapy of fibrotic CKD.

BACKGROUND The impact of occult HBV infection with HBsAg-/HBV DNA+ (OBI) on severity of liver fibrosis remains unclear. METHODS A total of 1772 HBsAg negative but anti-HBc positive subjects stratified between OBI and non-OBI were selected for long-term carriage leading to at least two of four liver fibrosis indexes elevated: hyaluronic acid (HA), laminin (LN), type III procollagen peptide (PCIII) or type IV collagen (CIV) tested in a Chinese hospital. Patients were tested for serum viral load, and HBV markers and histopathological changes in liver biopsies. RESULTS OBI was identified in 148 liver fibrosis patients (8.4%), who had significantly higher levels of HA, LN, PCIII and CIV than 1624 fibrotic patients without OBI (P<0.05). In 36 OBI patients with liver biopsy, significant correlations were observed between OBI viral load and serum HA levels (P=0.0148), PCIII levels (P=0.0106) and pathological HAI scores (P<0.0001), HAI scores and PCIII levels (P=0.0383), HBcAg immunohistochemical (HBcAg-IHC) scores and HA levels (P=0.0003), and HBcAg-IHC scores and PCIII levels (P=0.0303), respectively. Positive FISH was significantly more frequent in OBIs (80.6% versus 37.5%). HBcAg was detected in liver tissues of 52.8% and HBsAg in 5.6% of OBI patients. CONCLUSIONS OBI status appears associated with liver fibrosis severity.

Novel adenovirus vectors are an important antigen delivery platform for vaccine development. Understanding the immune diversity between different adenoviral vectors is critical to design the proper vaccine against an aim disease.

Recombinant adenovirus vector has been widely used in vaccine development. Due to the pre-existing immunity of human adenovirus type 5 (HAd5) in humans, a range of rare human and chimpanzee adenovirus vectors have been developed. In the previous study, we constructed novel adenovirus vector Sad23L and Ad49L based on simian adenovirus type 23 (SAd23) and human adenovirus type 49 (HAd49), which were used in the development of ZIKV and COVID-19 vaccines. However, the levels of pre-existing neutralizing antibody (NAb) of HAd49 and SAd23 remain unclear in China. In this study, we measured NAbs titers of HAd5, HAd49, and SAd23 in 600 healthy blood donors from 6 regions across China. NAb titer of HAd49 or SAd23 was significantly lower than that of HAd5 (p < 0.001). There was no significant difference in seroprevalence and NAb titers of three adenoviruses between male and female donors. The seropositive rates of HAd5 and SAd23 increased with age growth in a positive correlation (p < 0.01), while in contrast to HAd5, HAd49, and SAd23 had a low level of pre-existing immunity in Chinese population, which suggested that Ad49L and Sad23L vectors could be used in vaccine development for humans.

Cats are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and risk transmitting viruses to naive cats or humans. Here, based on our novel adenovirus-vectored COVID-19 vaccine, the immunogenicity of Sad23L-nCoV-S vaccine was evaluated in cats by prime-boost vaccinations. Five cats were primed with a dose of 108 plaque-forming units (PFUs) Sad23L-nCoV-S vaccine and then boosted with an equal dose of same vaccine at a 4-week interval. Cat serum neutralizing antibody (NAb) titers (the sample dilution at which 50% inhibitory concentration [IC50]) were measured as IC50 15,849 to wild-type strain, IC50 6,591 to Alpha, IC50 2,315 to Beta, IC50 2,744 to Gamma, IC50 1,848 to Delta, and IC50 318 to Omicron variants of pseudotyped SARS-CoV-2 viruses at week 6 post-prime vaccination. All NAb levels to these five variants were ≥IC50 49 from vaccinated cats at week 10, while 48.8% to Delta and 100% to Omicron variants were

To control the coronavirus disease 2019 (COVID-19) pandemic, there is an urgent need for simple, rapid, and reliable detection methods to identify severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, especially in community hospitals or clinical centers. The SARSCoV-2 nucleocapsid protein (NP) is an important index for diagnosis of COVID19. Here, we proposed a smartphone-based high-throughput fiber-integrated immunosensing system (HFIS) for detecting the SARS-CoV-2 NP in serum samples within 45 min. For the testing of NP standards, the linear detection range was 7.8-1000 pg/mL, the limit of detection was 7.5 pg/mL, and the cut-off value was 8.923 pg/mL. Twenty-five serum samples from clinically diagnosed COVID19 patients and 100 negative control samples from healthy blood donors were tested for SARS-CoV-2 NP by HFIS, and the obtained results were compared with those of ELISA and Simple Western analysis. The results showed that the HFIS sensitivity and specificity were 72% [95% confidence interval (CI): 52.42-85.72%] and 100% (95% CI: 96.11-100%), respectively, which significantly correlated with those from the commercial ELISA kit and Simple Western analysis. This portable high-throughput HFIS assay could be an alternative test for detecting SARS-CoV-2 NP in blood samples on site.

Brucellosis is a worldwide infectious zoonotic disease, posing severe threats to human health and social-economic development. By comparing with time-consuming, low sensitive and non-quantitative conventional serological methods, herein, protein G (prG) coupled with europium nanospheres (EuNPs) (detection probe) and highly purified Brucella lipopolysaccharide (LPS) (capture antigen) were used to develop a novel time-resolved fluorescence lateral flow immunoassay (TF-LFIA) for detecting anti-Brucella IgG antibody in human plasmas. The entire testing took 15 min. With a satisfactory purity, the purified LPS weakly cross-reacted with Y. enterocolitica O9 diagnostic antibody; however, none reacted with sera from patients with other Gram-negative bacterial infections. Following coefficient of determination (R-2 = 0.9961), 0.3 IU/mL was reported as the limit of detection (LOD), much lower than those of Serological Agglutination Test (SAT), Rose-Bengal Plate Agglutination Test (RBPT) and colloidal gold LFIA (CG-LFIA). Intra-day and inter-day precisions (CV, coefficient variation) of TF-LFIA varied less than 8% or 12 %, while intra-day and inter-day accuracies were 94-106 % or 93-107 %, respectively. The correlation coefficient (R-2) of TF-LFIA measurement to the different concentrations of spiked Brucella antibody was 0.9967, suggesting TF-LFIA had high reliability and reproducibility. TF-LFIA was demonstrated for 100 % specificity, 98.57 % sensitivity and 99.63 % accuracy in detection of Brucella antibody from clinical samples, respectively, significantly higher compared to SAT and RBPT. In conclusion, the established TF-LFIA is a simple, rapid and quantitative immunoassay for early diagnosis or epidemiological surveillance of Brucella infection in humans. (C) 2021 Elsevier B.V. All rights reserved.

The level of neutralizing antibody (NAb) to SARS-CoV-2 could be used to evaluate the acquired protective immunity of COVID-19 patients or vaccinees. Here we develop a track-etched microporous membrane filtration microplate (TEM) and optical fibers transmitted immunosensing smartphone platform (TEMFIS) based surrogate virus neutralization test (TEMFIS-sVNT) for rapid one-step testing of NAb to SARS-CoV-2. Coefficient variation (CV) of intra-assay and inter-assay precisions of TEMFIS-sVNT varied below 9% or 14%, respectively. By agreement with pseudovirus neutralization test (pVNT) and ELISA-sVNT for testing of serum samples from 41 COVID-19 patients, 50 COVID-19 vaccinees and 320 healthy blood donors (P = 0.895), TEMFIS-sVNT detected the NAb positivity (sensitivity) in 92.68% COVID-19 patients and 76% vaccinees, but the NAb negativity (specificity) in 100% blood donors. In conclusion, TEMFIS-sVNT can be used for quantitatively point-of-care testing of neutralizing antibody to SARS-CoV-2 in blood samples from COVID-19 patients and vaccinees.