Antibiotic-enzyme-inorganic nanoflowers based immunoassay for the ultrasensitive detection of Staphylococcus aureus.

In this work, we constructed a moderate and convenient approach for the determination of staphylococcus aureus (S. aureus) by using organic-inorganic flower-like hybrid nanoflowers and Pig IgG together in an enzyme-linked immunosorbent assay (ELISA) system. To ensure efficient capture, the hybrid nanoflowers were prepared by encapsulating horseradish peroxidase (HRP) and vancomycin (VAN) in the inorganic nanocrystal composites (calcium ion solution), just like the mimic biomineralization process. Owing to the self-assembly technique, the synthesized VAN-HRP-CaHPO4 nanoflowers (NFs) can not only retain the ability to particularly capture the gram-positive bacteria but also enhance the stability and enzymatic activity to achieve the signal output amplification. Then, taking advantage of the integration of signal amplification elements (HRP) and biorecognition unit (VAN), the VAN-HRP-CaHPO4 NFs were utilized as a new kind of capture & signal regent in the procedure of S. aureus detection. Based on this ELISA system, S. aureus could be clearly detected within the concentration ranging from 1.0 × 102 to 1.0 × 107 CFU mL-1. The detection limit was defined as 4.3 CFU mL-1, which performance is superior to some commercial ELISA kits. Additionally, this system detected the S. aureus in food samples and showed an acceptable recovery. As a cost-effective and sensitive platform, this proposed assay was enable to fulfill the requirement of a quick and effective detection of S. aureus.

LIS1, a glyco-humanized swine polyclonal anti-lymphocyte globulin, as a novel induction treatment in solid organ transplantation.

Anti-thymocyte or anti-lymphocyte globulins (ATGs/ALGs) are immunosuppressive drugs used in induction therapies to prevent acute rejection in solid organ transplantation. Because animal-derived, ATGs/ALGs contain highly immunogenic carbohydrate xenoantigens eliciting antibodies that are associated with subclinical inflammatory events, possibly impacting long-term graft survival. Their strong and long-lasting lymphodepleting activity also increases the risk for infections. We investigated here the in vitro and in vivo activity of LIS1, a glyco-humanized ALG (GH-ALG) produced in pigs knocked out for the two major xeno-antigens αGal and Neu5Gc. It differs from other ATGs/ALGs by its mechanism of action excluding antibody-dependent cell-mediated cytotoxicity and being restricted to complement-mediated cytotoxicity, phagocyte-mediated cytotoxicity, apoptosis and antigen masking, resulting in profound inhibition of T-cell alloreactivity in mixed leucocyte reactions. Preclinical evaluation in non-human primates showed that GH-ALG dramatically reduced CD4+ (p=0.0005,***), CD8+ effector T cells (p=0.0002,***) or myeloid cells (p=0.0007,***) but not T-reg (p=0.65, ns) or B cells (p=0.65, ns). Compared with rabbit ATG, GH-ALG induced transient depletion (less than one week) of target T cells in the peripheral blood (<100 lymphocytes/L) but was equivalent in preventing allograft rejection in a skin allograft model. The novel therapeutic modality of GH-ALG might present advantages in induction treatment during organ transplantation by shortening the T-cell depletion period while maintaining adequate immunosuppression and reducing immunogenicity.

Antibiotic and mammal IgG based lateral flow assay for simple and sensitive detection of Staphylococcus aureus.

Lateral flow assay (LFA), performed with simple devices and short detection time, is popular in field applications. Herein, a novel sandwich type-based LFA was constructed for high sensitivity and selectivity detection of Staphylococcus aureus (S. aureus). Vancomycin-immobilized gold nanoparticles (VAN-Au NPs) were utilized as the first identifier to capture S. aureus and the specificity was guaranteed by the second recognition agent of pig immunoglobulin G (IgG). In addition, gold growth was adopted for signal amplification to further improve the detection sensitivity. S. aureus could be directly assayed by this LFA within the concentration range of 1.0 × 103-1.0 × 108 cfu mL-1 with a detection limit of 1.0 × 103 cfu mL-1. Furthermore, the novel sandwich LFA realized S. aureus detection in food samples with admissible recoveries and established a rapid, simple, cost-effective and sensitive platform, could meet the demand for on-site testing of S. aureus.

Streptavidin-exposed magnetic nanoparticles for lectin magnetic separation (LMS) of Staphylococcus aureus prior to three quantification strategies.

A lectin magnetic separation (LMS) method for Staphylococcus aureus (S. aureus) was developed with the aim to improve the efficiency of magnetic nanoparticles and to expand the scope of bacterial recognition. Poly(ethylene glycol) (PEG)-mediated magnetic nanoparticles modified with streptavidin (MNP-PEG-SA) were synthesized and then applied to a two-step LMS based on the use of wheat germ agglutinin (WGA). Three specific methods for S. aureus detection (suitable for different requirements including detection time and sensitivity) were designed. The new LMS has improved anchoring efficiency (compared to two-step LMS methods) and requires a reduced number of magnetic particles. The Baird-Parker (B-P) method can detect S. aureus with a detection limit of 3 × 100 CFU·mL-1 within 15 h; the polymerase chain reaction (PCR) method can be finished within 4 h, with the lowest detection limit (LOD) of 3 × 102 CFU·mL-1. The LOD of HRP-pig IgG-based colorimetric method is 3 × 105 CFU·mL-1, and the method only lasts for 2 h. If combined with specific detection methods, it meets different needs for rapid detection of S. aureus. Graphical abstractSchematic representation of lectin magnetic separation (LMS) based on biotin-wheat germ agglutinin (WGA) and poly (ethylene glycol) (PEG)-mediated streptavidin-modified magnetic nanoparticles (MNP-PEG-SA) and three different quantification strategies (including B-P culture assay, PCR assay, and colorimetric assay) for S. aureus.

[Vancomycin-based fluorescent enzyme-linked immunoabsorbent assay for detection of Staphylococcus aureus].

In the study, fluorescent enzyme-linked immnoabsorbent assay for detection of Staphylococcus aureus was established with IgG from pig as capture antibody and quantum dot nanobeads (QBs) labeled vancomycin (QB-Vans) as testing antibody. Quantum dot of about 100 nm partical size nanobeads were prepared and linked with vancomycin. The optimum concentrations of salt ions were 0.01 mol/L, and the optimum pH was 6.0. Under the optimum conditions, the detection sensitivity for S. aureus was 104 CFU/mL, and there was no cross-reaction with other pathogenic bacteria. Thus, the method could be used for rapid screening of S. aureus, for the clinical monitoring and foodborne pathogens detection.

Gold nanostar-enhanced surface plasmon resonance biosensor based on carboxyl-functionalized graphene oxide.

A new high-sensitivity surface plasmon resonance (SPR) biosensor based on biofunctional gold nanostars (AuNSs) and carboxyl-functionalized graphene oxide (cGO) sheets was described. Compared with spherical gold nanoparticles (AuNPs), the anisotropic structure of AuNSs, which concentrates the electric charge density on its sharp tips, could enhance the local electromagnetic field and the electronic coupling effect significantly. cGO was obtained by a diazonium reaction of graphene oxide (GO) with 4-aminobenzoic acid. Compared with GO, cGO could immobilize more antibodies due to the abundant carboxylic groups on its surface. Testing results show that there are fairly large improvements in the analytical performance of the SPR biosensor using cGO/AuNSs-antigen conjugate, and the detection limit of the proposed biosensor is 0.0375 µg mL(-1), which is 32 times lower than that of graphene oxide-based biosensor.