Immunomodulation of nutritional formula containing epigallocatechin-3-gallate, ginseng extract, and polydextrose on inflammation and macrophage polarization.

Single nutrient likes polyphenol or dietary fiber have been exhaustively investigated to validate their positive intervention in health or disease. Meanwhile, the common interaction of inner systems with the nutrient complex has not been well elucidated, which raises the scientific issue of the modulatory effect of the nutrient complex on immunity. The representative prebiotics of epigallocatechin-3-gallate (EGCG), ginseng extract, and polydextrose (PDX) were selected on behalf of the classification of polyphenol, flavone or polysaccharides, and dietary fiber to generally cover the daily food intake in this study to explore their intervention in inflammation and macrophage polarization. The intervention of selected nutrients on inflammation and macrophage polarization has been evaluated against macrophages to unveil their comprehensive effects. The synergistic effect of selected nutrients was demonstrated by inhibiting M1 macrophage polarization and the promotion of M2 macrophage polarization. Then, the nutrient formula was set up to verify the intervention effect, and the results revealed the significant inhibition of cell inflammation and the effect on cell proliferation through promoting the cell cycle in the G2 phase. The nutrient complex could inhibit M1 macrophage polarization to inhibit M1-mediated inflammation and promote M2 macrophages for anti-inflammatory effect and enhance cell phagocytosis. Moreover, the varied intervention effects of the nutrient complex with different formulas could be summarized. In general, the formula containing EGCG, ginseng extract, and PDX was demonstrated to possess an enhanced immunomodulatory effect on cell inflammation and macrophage polarization, which could potentially inspire the investigation of complex nutrients in health and diseases.

Digestibility of Malondialdehyde-Induced Dietary Advanced Lipoxidation End Products and Their Effects on Hepatic Lipid Accumulation in Mice.

Advanced lipoxidation end products (ALEs) are formed by modifying proteins with lipid oxidation products. The health effects of ALEs formed in vivo have been extensively studied. However, the digestibility, safety, and health risk of ALEs in heat-processed foods remain unclear. This investigation was performed to determine the structure, digestibility, and effect on the mice liver of dietary ALEs. The results showed that malondialdehyde (MDA) was able to alter the structure of myofibrillar proteins (MPs) to form linear, loop, and cross-linked types of Schiff bases and dihydropyridine derivatives under simulated heat processing, leading to the intra- and intermolecular aggregation of MPs and, thus, reducing the digestibility of MPs. In addition, dietary ALE intake resulted in abnormal liver function and lipid accumulation in mice. The core reason for these adverse effects was the destructive effect of ALEs on the intestinal barrier. Because the damage to the intestinal barrier leads to an increase in lipopolysaccharide levels in the liver, it induces liver damage by modulating hepatic lipid metabolism.

Nanobody-Based Electrochemical Immunoassay for Sensitive Detection of Peanut Allergen Ara h 1.

Peanut is widely used for food supplementation with potential allergic reactions in infants and adults, which prompted the development of reliable and accurate detection of peanut allergens with emphasis on Ara h 1. In this study, a nanobody (Nb)-based micro-total electrochemical immunoassay (Nb-µTEI) was proposed to be generated. Generally, an alpaca was immunized with Ara h 1 to yield a Nb reservoir for selection of four specific Nbs. Nb-mediated immunocapturing allowed the identification of the target as Ara h 1. The Nb-based electrochemical immunoassay was developed by constructing a capturing electrode with cycles of signal enhancement. After construction of the capturing electrode, Nb152 with HA-tag was directly applied to connect immobilized anti-HA IgG for the capture of different concentrations of Ara h 1, which was labeled by biotinylated Nb152 to facilitate signal development with alkaline phosphatase conjugated streptavidin (SA-ALP). A linear range from 4.5 to 55 ng/mL was acquired with LOD and LOQ of 0.86 and 2.10 ng/mL, respectively, with an 11-fold increase of the sensitivity compared with the established sandwich ELISA. The dedicated immunoassay was verified by detecting the antigen spiked in food samples and demonstrated the successful conjugation of Nb with advanced detecting techniques.

Detection of ß-lactoglobulin under different thermal-processing conditions by immunoassay based on nanobody and monoclonal antibody.

The problems of inaccurate detection values of thermal-processed ß-lactoglobulin (ß-LG) content seriously affect the screening of allergens. A monoclonal antibody (mAb) against ß-LG was successfully prepared and a highly sensitive sandwich ELISA (sELISA) was constructed with specific nanobody (Nb) as the capture antibody with detection limit of 0.24 ng/mL. Based on this sELISA, the ability of Nb and mAb to recognize ß-LG and ß-LG interacting with milk components was explored. Combined with protein structure analysis to elaborate the mechanism of shielding ß-LG antigen epitopes during thermal-processing, thus enabling the differentiation between pasteurized and ultra-high temperature sterilized milk, the detection of milk content in milk-containing beverages, and the highly sensitive detection and analysis of ß-LG allergens in dairy-free products. The method provides methodological support for identifying the quality of dairy products and reducing the risk of ß-LG contamination in dairy-free products.

A Plasmonic Fluor-Lightened Microneedle Array Enables Ultrasensitive Multitarget Whole Blood Diagnosis of Anemia in A Paper Origami-Based Device.

This work reports a portable, origami-type paper device with a plasmonic fluor-labeled microneedle sensing module for the multiplexed quantification of anemia biomarkers in whole blood. Sequential steps, including serum separation, target enrichment, and multiplexed readout by a gel imager, are rapidly accomplished with the flexible and highly integrated device. The microneedle array enabled efficient sampling of trace targets from ng mL-1 to pg mL-1 level. Combined with the plasmonic fluor label, the signal is improved by ≈7.6 folds compared with the flat substrate-based assay. The device is applied to simultaneously quantify hemoglobin (Hb), ferritin, folic acid (FA), and vitamin B12 (VB12 ), which are four anemia biomarkers distributed in different environments with different concentration ranges. Featured by the small sample volume (150 µL), short assay time (20 min), low cost (2 $), robust stability, and user-friendliness, the device is promising for the rapid and accurate diagnosis of anemia in real practice.

Gut Microbiota-Derived Indole Derivatives Alleviate Neurodegeneration in Aging through Activating GPR30/AMPK/SIRT1 Pathway.

SCOPE: Tryptophan (Trp) metabolites are closely related to neurological diseases, whereas, the underlying mechanism related to the alleviative effects of Trp metabolites on neurodegeneration in aging remains unclear. This study aims to evaluate the protective effects and mechanisms of Trp metabolites on neurodegeneration in aging process. METHODS AND RESULTS: The neuroprotective properties of Trp metabolites are evaluated in vitro and in vivo experimental model. Trp metabolites such as indole, indole-3-acetic acid (IAA), indole-3-propionic acid (IPA), indole-3-lactic acid (ILA), and indole-3-carboxyaldehyde (Icld) could significantly reduce oxidative stress, inflammation, and neuronal apoptosis induced by H2 O2 in HT-22 cells. Meanwhile, indoles could upregulate the expressions of G protein-coupled receptor 30 (GPR30)/5'-adenosine monophosphate (AMP)-activated protein kinase (AMPK)/silent information regulator 1 (SIRT1) pathway in vitro. Furthermore, the neuroprotective effects of IAA and IPA are unveiled through activation of GPR30/AMPK/SIRT1 pathway in d-galactose induced aging mice. Finally, the regulatory effects of indoles on GPR30/AMPK/SIRT1 pathway are further confirmed by pretreating HT-22 and Neuro-2a with GPR30 antagonist of G15. In that case, indoles are furtherly proved with inhibitory effects on neurodegeneration by activating the GPR30/AMPK/SIRT1 pathway in aging process. CONCLUSIONS: The findings reveal that Trp metabolites significantly improve neurodegeneration via GPR30/AMPK/SIRT1 pathway in aging process. This study provides the potential novel intervention strategy and target to prevent the neurodegeneration.

Exploration of Specific Nanobodies As Immunological Reagents to Detect Milk Allergen of ß-Lactoglobulin without Interference of Hydrolytic Peptides.

Milk proteins are widely used for food supplementation, despite the potential risk of food allergy, especially against ß-lactoglobulin (BLG), which makes BLG surveillance critical. Possible interaction of detecting antibodies with BLG-derived peptides will result in unprecise inspection. Thus, in this study, it was proposed to generate nanobodies (Nbs) and validate the immunological detection of intact BLG rather than hydrolytic peptides. Nbs were successfully retrieved and characterized with high stability and target specificity. A competitive enzyme-linked immunosorbent assay (cELISA) was developed with a linear range from 39 to 10,000 ng/mL and a detection limit (LOD) of 4.55 ng/mL, with a recovery of 86.30%-95.09% revealed by analysis of spiked samples. Meanwhile, a sandwich ELISA (sELISA) was established with Nb82 and BLG polyclonal antibody (pAb-BLG) providing a linear range from 29.7 to 1250 ng/mL and an LOD of 13.82 ng/mL with a recovery of 87.82%-103.97%. The interaction of selected Nbs with BLG-derived peptides was investigated by Nb structure modeling and BLG docking. No binding on hydrolytic peptides was revealed, confirming the precision of Nb-mediated immunoassays. In summary, this study successfully identified BLG-specific Nbs for immunoassay development and guaranteed the monitoring of intact BLG without interference of hydrolytic peptides, providing experimental evidence that our Nbs recognize intact food allergen.

Identification of Serum Ferritin-Specific Nanobodies and Development towards a Diagnostic Immunoassay.

Serum ferritin (SF) is an iron-rich protein tightly connected with iron homeostasis, and the variations are frequently observed in diseased states, including iron-deficiency anemia, inflammation, liver disease, and tumors, which renders SF level an indicator of potential malignancies in clinical practice. Nanobodies (Nbs) have been widely explored and developed into theranostic reagents. Surprisingly, no reports stated the identification of anti-SF Nbs, nor the potential of such Nbs as a diagnostic tool. In this study, we generated SF-specific Nbs and provided novel clinical diagnostic approaches to develop an immunoassay. An immune library was constructed after immunizing an alpaca with SF, and five Nbs specifically targeting human SF were retrieved. The obtained Nbs exhibited robust properties including high stability, affinity, and specificity. Then, an ELISA-based test using a heterologous Nb-pair was developed. The calibration curve demonstrated a linear range of SF between 9.0 to 1100 ng/mL, and a limit of detection (LOD) of 1.01 ng/mL. The detecting recovery and coefficient variation (CV) were determined by spiking different concentrations of SF into the serum sample, to verify the successful application of our selected Nbs for SF monitoring. In general, this study generated SF-specific Nbs and demonstrated their potential as diagnostic immunoassay tools.

Identification and Characterization of Specific Nanobodies against Trop-2 for Tumor Targeting.

Trophoblast cell-surface antigen 2 (Trop-2) is a tumor-associated antigen that is connected with the development of various tumors and has been identified as a promising target for tumor immunotherapy. To date, the immunotherapy against Trop-2 mainly relies on the specific targeting by monoclonal antibody in antibody-drug conjugate (ADC). Alternatively, the single domain antibodies of nanobodies (Nbs) possesses unique properties such as smaller size, better tissue penetration, etc., to make them good candidates for tumor targeting. Thus, it was proposed to develop anti-Trop-2 Nbs for tumor targeting in this study. Generally, three consecutive rounds of bio-panning were performed against immobilized recombinant Trop-2, and yielded three Nbs (Nb60, Nb65, and Nb108). The affinity of selected Nbs was determined in the nanomolar range, especially the good properties of Nb60 were verified as a promising candidate for tumor labeling. The binding to native Trop-2 was confirmed by flow cytometry against tumor cells. The inhibitory effects of the selected Nbs on tumor cell proliferation and migration were confirmed by wound healing and Transwell assay. The clear localization of the selected Nbs on the surface of tumor cells verified the potent labeling efficiency. In conclusion, this study provided several Nbs with the potential to be developed as targeting moiety of drug conjugates.

Different Dose of Sucrose Consumption Divergently Influences Gut Microbiota and PPAR-γ/MAPK/NF-κB Pathway in DSS-Induced Colitis Mice.

Sugar reduction and sugar control are advocated and gaining popularity around the world. Sucrose, as the widely consumed ingredient in our daily diet, has been reported a relation to gastrointestinal diseases. However, the role of sucrose in inflammatory bowel disease remains controversial. Hence, our study aimed to elucidate the potential role of three doses of sucrose on DSS-induced colitis in C57BL/6 mice and the underlying mechanisms. The results showed that low-dose sucrose intervention alleviated colitis in mice, reducing the expression of inflammatory cytokines and repairing mucosal damages. In contrast, high-dose sucrose intervention exacerbated colitis. Furthermore, three doses of sucrose administration markedly altered gut microbiota composition. Notably, the low-dose sucrose restored microbial dysfunction and enhanced the production of short chain fatty acids (SCFAs). Specifically, the abundance of SCFAs-producing bacteria Faecalibaculum, Bacteroides, and Romboutsia were increased significantly in the LOW group. Consistently, PPAR-γ, activated by SCFAs, was elevated in the LOW group, thereby inhibiting the MAPK/NF-κB pathway. Together, our study demonstrates the differential effects of sucrose on colitis at different doses, providing a scientific basis for measuring and modifying the safe intake level of sugar and providing favorable evidence for implementing sugar reduction policies.

Versatile Application of Nanobodies for Food Allergen Detection and Allergy Immunotherapy.

The unique characteristics of camelid heavy-chain only antibody (HCAb) derived nanobodies (Nbs) have facilitated their employment as tools for research and application in extensive fields including food safety inspection, diagnosis and therapy of diseases, etc., to develop immune detecting techniques or alternative candidates of conventional antibodies as diagnostic and therapeutic reagents. The wide application in the fields of food allergen inspection and immunotherapy has not been addressed as not much results published in the literature. The robust properties and straightforward selecting strategy of Nbs impel the advantageous employment compared with monoclonal antibodies (mAbs) to establish immunoassay and serve as blocking antibodies to compete immunoglobulin E (IgE) binding epitopes on food allergens. More and more efforts have been invested to develop specific Nbs against food allergen proteins, such as macadamia allergen of Mac i 1, peanut allergen of Ara h 3, and lupine allergen of Lup an 1, which demonstrated the potential of Nbs for research and application in food allergen surveillance. Meanwhile, the paratopes of Nbs preferably targeting the unique epitopes of food allergens can provide more possibilities to serve as blocking antibodies to shield IgE binding epitopes for food allergy immunotherapy. Regardless, the research and application of Nbs in the field of food allergen and allergic reactions are expected to attract dramatic focus and produce promising research outputs.

Sweeteners Maintain Epithelial Barrier Function Through the miR-15b/RECK/MMP-9 Axis, Remodel Microbial Homeostasis, and Attenuate Dextran Sodium Sulfate-Induced Colitis in Mice.

Non-nutritive sweeteners are the most widely used food additives designed to provide sweetness and reduce caloric intake. Studies have confirmed a link between sweeteners and colitis, yet supporting scientific data remain exiguous and controversial. In this study, three common sweeteners (Saccharin sodium, Stevioside, and Sucralose) in acceptable daily intake dosage were added to water in order to determine their effects on dextran sodium sulfate-induced colitis in mice. Our results show that the three sweeteners meliorate colitis to varying degrees─Saccharin exerts the most pronounced effect, followed by Stevioside and Sucralose. Intake of sweeteners alleviates colitis symptoms, alters gut microbiota, reshapes the TH17/Treg balance, protects the intestinal barrier, and reduces inflammation. Most significantly, sweeteners can enhance the abundance of Mucispirillum and Alistipes, which are conducive to colitis recovery, and upregulate the expression of E-cadherin through the miR-15b/RECK/MMP-9 axis to improve intestinal barrier integrity. Moreover, by inhibiting the MMP-9/AKT/NF-κB pathway, inflammation is relieved, as reflected in the restoration of the Th17/Treg balance. Our results link the consumption of sweeteners to the remission of colitis, which provides new scientific evidence for the safe use of sweeteners.

Intrabody Targeting HIF-1α Mediates Transcriptional Downregulation of Target Genes Related to Solid Tumors.

Uncontrolled growth of solid tumors will result in a hallmark hypoxic condition, whereby the key transcriptional regulator of hypoxia inducible factor-1α (HIF-1α) will be stabilized to activate the transcription of target genes that are responsible for the metabolism, proliferation, and metastasis of tumor cells. Targeting and inhibiting the transcriptional activity of HIF-1 may provide an interesting strategy for cancer therapy. In the present study, an immune library and a synthetic library were constructed for the phage display selection of Nbs against recombinant PAS B domain protein (rPasB) of HIF-1α. After panning and screening, seven different nanobodies (Nbs) were selected, of which five were confirmed via immunoprecipitation to target the native HIF-1α subunit. The inhibitory effect of the selected Nbs on HIF-1 induced activation of target genes has been evaluated after intracellular expression of these Nbs in HeLa cells. The dramatic inhibition of both intrabody formats on the expression of HIF-1-related target genes has been confirmed, which indicated the inhibitory efficacy of selected Nbs on the transcriptional activity of HIF-1.

Selection of Specific Nanobodies against Lupine Allergen Lup an 1 for Immunoassay Development.

The declaration of lupine supplements is mandatory to avoid lupine allergy for sensitive individuals. However, reliable detection methods against lupine allergen remain critical to prevent the unintended consumption of allergen contaminated food. In this study, we have immunized an alpaca with lupine protein extracts and retrieved nanobodies (Nbs). Nevertheless, the target antigen has been recognized as Lup an 1, which has been classified as ß-conglutin, and confirmed to connect with lupine allergy. After selection of the best Nb-pair, a sandwich enzyme-linked immunosorbent assay (ELISA) has been developed providing a linear range of 0.036-4.4 µg/mL with detection limit of 1.15 ng/mL. This immunoassay was confirmed by detecting the samples with spiked allergen, and a recovery from 86.25% to 108.45% with coefficient of variation (CV) less than 4.0% has been determined. Generally, this study demonstrated the selection of Nbs against allergen with crude protein content to develop the immunoassay for lupine surveillance in foods.

Dose-Dependent Beneficial Effects of Tryptophan and Its Derived Metabolites on Akkermansia In Vitro: A Preliminary Prospective Study.

Akkermansia muciniphila, a potential probiotic, has been proven to lessen the effects of several diseases. As established, the relative abundance of Akkermansia is positively correlated with tryptophan metabolism. However, the reciprocal interaction between tryptophan and Akkemansia is still unclear. Herein, for the first time, the possible effects of tryptophan and its derived metabolites on A. muciniphila were preliminarily investigated, including growth, physiological function, and metabolism. Obtained results suggested that 0.4 g/L of tryptophan treatment could significantly promote the growth of A. muciniphila. Notably, when grown in BHI with 0.8 g/L of tryptophan, the hydrophobicity and adhesion of A. muciniphila were significantly improved, potentially due to the increase in the rate of cell division. Furthermore, A. muciniphila metabolized tryptophan to indole, indole-3-acetic acid, indole-3-carboxaldehyde, and indole-3-lactic acid. Indoles produced by gut microbiota could significantly promote the growth of A. muciniphila. These results could provide a valuable reference for future research on the relationship between tryptophan metabolism and A. muciniphila.

Promotion Effect of EGCG on the Raised Expression of IL-23 through the Signaling of STAT3-BATF2-c-JUN/ATF2.

Tea polyphenol of epigallocatechin-3-gallate (EGCG) has been verified to possess multiple biological activities. Interleukin-23 (IL-23) is a heterodimeric cytokine consisting of two subunits of IL-23p19 and IL-12p40, with the functionality in regulating the production of cytokines under physiological or pathological conditions. By serendipity, the raised expression of IL-23 was observed after treating cells with EGCG, whereas the detailed mechanism remains poorly understood. This study was proposed to investigate the signaling related to EGCG-induced IL-23. The raised expression of IL-23 was confirmed primarily by intraperitoneally injecting with different concentrations of EGCG (0, 20, 50, 80 mg/kg) into BALB/c mice, and the raised expression was confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. Results from enzyme-linked immunosorbent assay (ELISA) revealed the increase of IL-23 in serum from 116.09 to 153.90 pg/mL after treating with EGCG. The same results were also observed in RAW264.7 and peritoneal macrophages after treating with EGCG (0, 1, 5, 10, 25 µM) with the increased tendency of IL-23 in cultural medium (7.98 to 25.38 pg/mL for RAW264.7; 3.64 to 260.93 pg/mL for peritoneal macrophages). After preliminary exploration of the signaling related to the increased IL-23, the classical signaling pathways and key transcription factors, such as nuclear factor kappa-B (NF-κB), mitogen-activated protein kinase (MAPK) signaling pathways, and interferon regulatory factor 5 (IRF5), were demonstrated with no relevant contribution. A further study revealed the involvement of the key transcription factor of BATF2, which could antagonistically modulate the transcription and translation of IL-23. The signaling of STAT3-BATF2-c-JUN/ATF2-IL-23 has been further verified in RAW264.7 macrophages using the STAT3 inhibitor of AG490 and the activator of Colivelin TFA. The results indicated that EGCG inhibits the phosphorylation of STAT3 to facilitate the decreased level of BATF2, which contributed to the increased level of IL-23 by the enhancing heterodimerization of c-JUN and ATF2.

Function Characterization of Endogenous Plasmids in Cronobacter sakazakii and Identification of p-Coumaric Acid as Plasmid-Curing Agent.

Virulence traits and antibiotic resistance are frequently provided by genes located on plasmids. However, experimental verification of the functions of these genes is often lacking due to a lack of related experimental technology. In the present study, an integrated suicide vector was used to efficiently and specifically delete a bacterial endogenous plasmid in Cronobacter sakazakii. The pESA3 plasmid was removed from C. sakazakii BAA-894, and we confirmed that this plasmid contributes to the invasion and virulence of this strain. In addition, the pGW1 plasmid was expunged from C. sakazakii GZcsf-1, and we confirmed that this plasmid confers multidrug resistance. We further screened plasmid-curing agents and found that p-coumaric acid had a remarkable effect on the curing of pESA3 and pGW1 at sub-inhibitory concentrations. Our study investigated the contribution of endogenous plasmids pESA3 and pGW1 by constructing plasmid-cured strains using suicide vectors and suggested that p-coumaric acid can be a safe and effective plasmid-curing agent for C. sakazakii.

Unbiased Immunization Strategy Yielding Specific Nanobodies against Macadamia Allergen of Vicilin-like Protein for Immunoassay Development.

Macadamia nut contains important food allergens that potentially cause allergic reactions with severe adverse effects in infants and adults. Reliable and accurate detection of macadamia is critical to avoid allergic reactions. However, knowledge on macadamia allergen is scarce and a reliable detection method has not been reported, yet. In this study, an unbiased immunization and selection strategy was employed to select nanobodies (Nbs) recognizing specifically macadamia allergen, as well as to establish a detection method to unveil a macadamia protein contamination. An alpaca was immunized with a crude protein extract of macadamia followed by construction of a Nb library from its lymphocytes. The panning and screening of this immune Nb repertoire resulted in the selection of six target-specific Nbs. Nb-mediated immuno-capturing combined with mass spectrometry allowed us to identify the target as the macadamia vicilin-like antimicrobial peptides 2-3 (MiAMP2), a novel food allergenic protein abbreviated as Mac i 1. Later on, an immunoassay of a heterologous sandwich ELISA method based on the selected Nb-pairs was established, providing a linear response in the range of 0.442-2,800 µg/mL and with a limit of detection of 27.1 ng/mL. The dedicated immunoassay has been verified by detecting the antigen spiked in food samples. Our study provided evidence for the successful application of the unprejudiced strategy to retrieve Nbs against a priori undefined macadamia allergen. These target-specific Nbs were used to design a highly reliable and effective immunoassay.

Ameliorative Effect of Dietary Tryptophan on Neurodegeneration and Inflammation in d-Galactose-Induced Aging Mice with the Potential Mechanism Relying on AMPK/SIRT1/PGC-1α Pathway and Gut Microbiota.

Dietary tryptophan affects intestinal homeostasis and neurogenesis, whereas the underlying mechanism and the reciprocal interaction between tryptophan and gut microbiota in aging are unclear. This investigation was performed to determine the effect and mechanism of tryptophan on intestinal- and neuro- health in aging. In present study, the 0.4% tryptophan diet significantly ameliorated the oxidative stress and inflammation in the aging mice, potentially through the regulation of 5'-adenosine monophosphate (AMP)-activated protein kinase (AMPK) and nuclear factor κB (NF-κB) pathways. The 0.4% tryptophan diet increased the levels of indoles in colon contents, which indicated the potential contribution of tryptophan metabolites. Microbiome analysis revealed that the 0.4% tryptophan diet raised the relative abundance of Akkermansia in aging. The ameliorated effect of 0.4% tryptophan on neurodegeneration and neuroinflammation was summarized to potentially rely on the brain-derived neurotrophic factor- (BDNF) and NF-κB-related pathways. These findings provide the research evidence for the beneficial effect of tryptophan on aging.

Selection of specific nanobodies to develop an immuno-assay detecting Staphylococcus aureus in milk.

The interaction between conventional immunoglobulins (Igs) and the Ig-binding surface proteins of Staphylococcus aureus (S. aureus) have obstructed the development of immuno-assays to detect these bacteria. The current study aimed to select nanobodies (Nbs) recognizing specifically S. aureus and to establish an immuno-assay to uncover S. aureus contaminations in foods. An alpaca was immunized with an inactivated S. aureus strain followed by the construction of a Nb library from which four target-specific Nbs were retrieved. Subsequently, a sandwich ELISA employing the Nb147 and biotinylated-Nb147 pair to capture and to detect S. aureus, respectively, was established to possess a detection limit of 1.4 × 105 colony forming units (CFU)/mL. The dedicated immuno-assay has been verified by detecting 10 CFU/mL of S. aureus in milk samples after an 8 h-enrichment step. This study provides the basis of an easy, reproducible and effective immuno-assay to screen for S. aureus contaminations in foods.