The bioluminescent luciferin-luciferase reaction is based on the oxidation of D-luciferin by oxygen in the presence of ATP and magnesium ions, catalyzed by firefly luciferase. The possibilities of using this reaction to study the influence of external effectors of a physical and chemical nature (temperature exposure, additions of drugs, membrane-active compounds, etc.) on living cells (prokaryotes and eukaryotes) are considered. Examples of the use of test systems based on living cells producing thermostable firefly luciferase for monitoring cellular homeostasis are given. The study of the kinetics of changes in the concentration of ATP and luciferase inside and outside cells made it possible to determine in dynamics the metabolic activity, cytotoxicity, and survival of cells under conditions of cellular stress, to study the processes of ATP synthesis/hydrolysis, and to evaluate the effectiveness of lytic agents in changing the permeability of the cell membrane.
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http://dx.doi.org/10.1007/s12551-022-00978-y | DOI Listing |
Vaccines (Basel)
December 2024
Shenzhen Neocurna Biotechnology Corporation, 12/F, Block B, Building 1, Yinxingzhijie Phase II, Longhua District, Shenzhen 518100, China.
The endosomal escape of lipid nanoparticles (LNPs) is crucial for efficient mRNA-based therapeutics. Here, we present a cationic polymeric micelle (cPM) as a safe and potent co-delivery system with enhanced endosomal escape capabilities. We synthesized a cationic and ampholytic di-block copolymer, poly (poly (ethylene glycol) methacrylate--hexyl methacrylate)--poly(butyl methacrylate--dimethylaminoethyl methacrylate--propyl acrylate) (p(PEGMA--HMA)--p(BMA--DMAEMA--PAA)), via reversible addition-fragmentation chain transfer polymerization.
View Article and Find Full Text PDFBiosensors (Basel)
January 2025
Laboratory of Biochemistry, Molecular Biology and Bioluminescent Systems Technology, Department of Physics, Chemistry and Mathematics, Federal University of Sao Carlos (UFSCAR), Rodovia João Leme dos Santos, km 110, Sorocaba 18052-780, SP, Brazil.
Firefly luciferases have been extensively used for bioanalytical applications, including their use as bioluminescent reporters, biosensors, and for bioimaging biological and pathological processes. Due to their intrinsic pH- sensitivity, in recent years we have demonstrated that firefly luciferases can also be harnessed as color- tuning sensors of intracellular pH. However, it is known that mammalian cells require temperatures higher than 36 °C, which red-shift the bioluminescence spectra of most firefly luciferases, decreasing their activities and the resolution of ratiometric pH analysis.
View Article and Find Full Text PDFCurr Microbiol
January 2025
Department of Plant Pathology, Pir Mehr Ali Shah Arid Agriculture University, Rawalpindi, Pakistan.
Xanthomonas citri pv. malvacearum (Xcm) associated with bacterial blight disease is a significant and widespread pathogen affecting cotton worldwide. The excessive use of harmful chemicals to control plant pathogens has exerted a negative impact on environmental safety.
View Article and Find Full Text PDFNat Chem
January 2025
State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, China.
The synthesis of large RNA with precise modifications at specific positions is in high demand for both basic research and therapeutic applications, but efficient methods are limited. Engineered DNA polymerases have recently emerged as attractive tools for RNA labelling, offering distinct advantages over conventional RNA polymerases. Here, through semi-rational designs, we engineered a DNA polymerase variant and used it to precisely incorporate a diverse range of modifications, including base modifications, 2'-ribose modifications and backbone modifications, into desired positions within RNA.
View Article and Find Full Text PDFJ Pediatr Surg
December 2024
Department of Surgery, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA. Electronic address:
Background: We sought to determine whether transamniotic stem cell therapy (TRASCET) could be a viable alternative for the fetal administration of genetically modified hematopoietic stem cells (HSCs) carrying a human hemoglobin subunit beta gene (hHBB) in a healthy syngeneic rat model.
Methods: Time-dated pregnant Lewis dams underwent volume-matched intra-amniotic injections in all their fetuses (n = 61) of a suspension of donor HSCs genetically modified with either both a hHBB gene and a firefly luciferase reporter gene (n = 42) or the firefly luciferase reporter gene alone to control for HBB-derived protein interspecies homology (n = 19) on gestational day 17 (E17; term = E21). Donor HSCs consisted of syngeneic cells phenotyped by flow cytometry with successful hHBB transduction confirmed by ELISA prior to administration in vivo.
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