Anti-FITC monoclonal antibody, clone 20

Fluorescence microscopy is one of the important instruments used in research and clinical laboratories on an everyday basis due to its wide range of applications ranging from a diagnostic tool to a molecular imaging device. However, the high cost, complex instrumentation, and requirement of sophisticated facilities limit its availability to school and college students, especially in developing and underdeveloped countries. In this work, we report the design and development of a slide-like portable total internal reflection fluorescence (TIRF) adapter, which can be integrated easily with Foldscope (a paper microscope) to observe and capture the fluorescence images using any smartphone. The adapter consists of a battery-poweredLED(light source), asmall piece of multimode optical fiber, a slide-like 3-D printed waveguide platform to hold the optical fiber and light emitting diode (LED) at an appropriate position. The potential of the adaptor is validated by capturing the fluorescence images of two most commonly used dyes i.e. fluorescein isothiocyanate (FITC) and rhodamine-B, after being immobilized on aminefunctionalized optical fiber probe. Further, this adaptor is successfully used to demonstrate (i) Forster resonance energy transfer phenomenon by employing FITC dye as donor and gold nanoparticles as quencher moiety and (ii) fluorescence bioassay by observing antigen-antibody interactions using human immunoglobulin G as the receptor and FITC-tagged goat anti-human immunoglobulin G as analyte. The inexpensive, portable and user-friendly TIRF adapter is a potential alternative to conventionally used fluorescence microscopes, especially for the school and college students and in resource-constrained clinical laboratories.

Background Non-small cell lung cancer (NSCLC) cells derived intracellular and extracellular programmed cell death ligand 1 (PD-L1) promoted cancer progression and drug resistance, and facilitated tumor immune evasion. However, the detailed molecular mechanisms are still largely unknown. In the present study, we aimed to explore the role of circular RNA circ-CPA4/let-7 miRNA/PD-L1 axis in the regulation of NSCLC progression, drug resistance and tumor immune microenvironment. Methods Real-Time qPCR and Western Blot analysis were conducted to examine gene expressions at transcriptional and translated levels, respectively. The regulatory mechanisms of circ-CPA4, let-7 miRNA and PD-L1 were validated by dual-luciferase reporter gene system and RNA pull-down assay. Cell growth and apoptosis were determined by CCK-8 assay, colony formation assay and Annexin V-FITC/PI double staining assay. Cell mobility was evaluated by transwell assay. Results Circ-CPA4 and PD-L1 were high-expressed, while let-7 miRNA was low-expressed in NSCLC cells and cancer tissues compared to the human bronchial epithelial (HBE) cells and their paired clinical normal adjacent tissues, respectively. Besides, knock-down of circ-CPA4 inhibited cell growth, mobility and epithelial-mesenchymal transition (EMT), and promoted cell death in NSCLC cells by downregulating PD-L1 through serving as a RNA sponge for let-7 miRNA. In addition, the NSCLC cells derived PD-L1-containing exosomes promoted cell stemness and increased resistance of NSCLC cells to cisplatin. Notably, by co-culturing the NSCLC cells with CD8(+)T cells isolated from human peripheral blood mononuclear cells (hPBMCs) in a transwell co-culturing system, we found that NSCLC cells inactivated CD8(+)T cells in a secreted PD-L1-dependent manner. Further results suggested that circ-CPA4 also positively regulated exosomal PD-L1, and the NSCLC cells with circ-CPA4 ablation re-activated CD8(+)T cells in the co-culturing system. Conclusion Taken together, circ-CPA4 regulated cell growth, mobility, stemness and drug resistance in NSCLC cells and inactivated CD8(+)T cells in the tumor immune microenvironment through let-7 miRNA/PD-L1 axis.