Quantitative analysis of autophagic flux by confocal pH-imaging of autophagic intermediates

Giuseppe Maulucci*, Michela Chiarpotto, Massimiliano Papi, Daniela Maria Samengo, Giovambattista Pani, Marco De Spirito

*Autore corrispondente per questo lavoro

Risultato della ricerca: Contributo in rivistaArticolo in rivistapeer review

37 Citazioni (Scopus)


Although numerous techniques have been developed to monitor autophagy and to probe its cellular functions, these methods cannot evaluate in sufficient detail the autophagy process, and suffer limitations from complex experimental setups and/or systematic errors. Here we developed a method to image, contextually, the number and pH of autophagic intermediates by using the probe mRFP-GFP-LC3B as a ratiometric pH sensor. This information is expressed functionally by AIPD, the pH distribution of the number of autophagic intermediates per cell. AIPD analysis reveals how intermediates are characterized by a continuous pH distribution, in the range 4.5-6.5, and therefore can be described by a more complex set of states rather than the usual biphasic one (autophagosomes and autolysosomes). AIPD shape and amplitude are sensitive to alterations in the autophagy pathway induced by drugs or environmental states, and allow a quantitative estimation of autophagic flux by retrieving the concentrations of autophagic intermediates.
Lingua originaleEnglish
pagine (da-a)1905-1916
Numero di pagine12
Stato di pubblicazionePubblicato - 2015


  • 3-methyladenine
  • LysoSensor
  • autophagic flux
  • autophagy
  • cathepsin B
  • chloroquine
  • fluorescence microscopy
  • mRFP-GFP-LC3
  • mechanistic target of rapamycin (serine/threonine kinase)
  • microtubule-associated protein 1 light chain 3
  • pH ratiometric sensor
  • pHrodo red
  • phosphatidylinositol 3-kinase catalytic subunit type 3
  • rapamycin
  • starvation


Entra nei temi di ricerca di 'Quantitative analysis of autophagic flux by confocal pH-imaging of autophagic intermediates'. Insieme formano una fingerprint unica.

Cita questo