We report a simple and ultra-sensitive surface enhanced Raman scattering (SERS) strip sensor based on silver nanoparticles (AgNPs) and lateral flow immunoassays (LFIAs). LFIAs are inexpensive, simple, portable and robust, thus making them commonplace in medicine, agriculture and food safety. However, their applications are limited due to the low signal intensity of the color-formation reaction based on the label accumulation. SERS is a powerful molecular spectroscopy technique for ultra-detection, which is based on the enhancement of the inelastic scattering from molecules located near nanostructured metallic surfaces when the molecules are illuminated and the surface plasmons are excited. Because of the rapidity and robustness of LFIAs and the high sensitivity of SERS, we introduce SERS into LFIAs (SERS-LFIA). Our SERS-LFIA demonstrates fast, excellent performance and is suitable for the semiquantitative examination of ultratrace analytes (Cr(3+)), with the limit of the detection (LOD) as low as 10(-5) ng mL(-1), which is 10(5)-fold more highly sensitive than those previously used to detect Cr(3+) within 15 min.
Using an enzyme-linked immunosorbent assay (ELISA) and limited dilution methods to screen and clone antigen-specific hybridoma cells is extremely time-consuming and labor-intensive. This work features a simple and rapid cell surface fluorescence immunosorbent assay (CSFIA), designed for the detection and isolation of antigen-specific hybridoma clones. In this assay, antigens are first anchored to the hybridoma cell surface through a dual-functioning molecular Oleyl-PEG4000-NHS. Specific antibodies secreted from hybridoma cells are then captured by the antigens on the cell surface. Positive hybridoma cells are stained using a fluorescently labeled anti-mouse IgG-Fc antibody. After the addition of a methylcellulose semisolid medium, positive clones are easily picked using a pipet. These positive cell clones can be used to produce monoclonal antibodies after direct expansion. Using this method, positive hybridoma clones against both malachite green and porcine epidemic diarrhea virus are selected with high efficiency. Compared to the ELISA-based method, the CSFIA-based method achieved the capability of isolating >2-fold more hybridoma clones in <25% of the corresponding processing time. In brief, the CSFIA-based method is highly efficient and inexpensive with a simple and direct operation, which is an excellent candidate method for antigen-specific positive clone isolation in a monoclonal antibody preparation.
Diabetic retinopathy (DR) is one of the more serious complications of diabetes. However, the mechanisms involved in DR are complex and still need to be investigated. The beneficial effects of fisetin have been widely reported, but whether it is beneficial in DR is not clear yet. This study was designed to investigate the regulatory role of fisetin in regulating DR and explore the involved mechanism. First, 30 mM glucose was used to establish DR cell model in vitro. Cell counting kit 8 (CCK8) assay was utilized to study the effects of fisetin on cell viability through treating human retinal microvascular endothelial cells (HRMECs) with 25, 50, and 100 μM fisetin. Transwell and wound healing assays were used to detect the function of fisetin on the migration and angiogenesis on HG-induced HRMECs. Finally, OE-VEGF was used as a mimic of VEGF, and western blotting (WB) was used to verify the targeting genes of fisetin. HG induced an increase in cell viability, cell migration, and angiogenesis in HRMECs, whereas fisetin inhibited these enhancements induced by HG through inhibiting VEGF. In conclusion, fisetin prevents angiogenesis in DR by downregulating VEGF.
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