Lentivirus Production for Autophagy Flux Detection

Autophagy is a highly conserved catabolic process in which long-lived cytoplasmic components or damaged organelles are sequestered by the formation of double-membraned autophagosomes. Mature autophagosomes ultimately fuse with lysosomes to form single-membraned autophagolysosomes that degrade or recycle their contents. Upon autophagy initiation, microtubule-associated protein 1 light chain 3 (LC3) is converted from LC3-I to the lipidated LC3-II and anchored to the autophagic membrane. The punctate distribution of LC3-II is regarded as a marker of autophagy induction, and it is closely related to the accumulation of autophagosomes. Lipidated LC3-II usually interacts with p62, which is a multifunctional protein that is degraded by the autophagic-lysosome pathway. The occurrence of complete autophagic flux is commonly reflected in the expression of LC3-II and p62. Currently, DNA constructs encoding fluorescent proteins fused to LC3 are widely employed for introduction into cells for monitoring autophagosome formation by fluorescence microscopy.

QVirus™ Platform has launched series of lentiviral packaging service of autophagy related biosensors, in which GFP and/or RFP tags are fused at the C-termini of the autophagosome marker LC3, allowing to detect the intensity of autophagy flux in real-time with more accuracy, clarity and intuitiveness. These biosensors provide an enhanced dissection of the maturation of the autophagosome to the autolysosome, which capitalizes on the pH difference between the acidic autolysosome and the neutral autophagosome. Our GFP-LC3, GFP-LC3 Control Mutant, and RFP-LC3 lentiviral particles provide bright fluorescence and precise localization of LC3 to the autophagosome, enabling live cell analysis of autophagy even in difficult-to-transfect cell types.

Features

  • Improved biosensor expression efficiency with lentiviral transduction
  • Ability to transfect dividing, non-dividing, and difficult-to-transfect cell types, such as primary cells or stem cells
  • Non-disruptive towards cellular function
  • Live cell time-lapse imaging of lentivirally-transduced cells
  • Enables visualization under different cell/disease states in live cell and in vitro analysis

QVirus™ Platform provides lentivirus production services to study the different stages of autophagy flux, making the autophagy study much easier. Our lentiviral biosensors enable convenient transduction of easy- and hard-to-transfect cell types with fluorescently-tagged proteins of interest. These packaged lentiviral particles provided higher efficiency of gene delivery and more homogeneous expression of introduced proteins compared to non-viral transfection methods. If you have any special requirements, please feel free to contact us.

Reference
1. Zhang Z , Singh R , Aschner M . Methods for the detection of autophagy in mammalian cells. 2016.

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