The majority of LC3 was diffusely localized in unstimulated (A) COS7 cells, whereas punctuated signals of LC3 signals of increase after induction of autophagy by vinblastine-stimulation for 2 hours (B). Cells were fixed in paraformaldehyd, followed by methanol. Permeabilization was performed with 0.3% Triton X-100 and endogenous LC3 subsequently detected by MAb 5F10

(Images by courtesy of I. Ciechomska and A. Tolkovsky, University of Cambridge, UK).

Immunoblot analysis of LC3 and Beclin using anti-LC3- and anti-Beclin monoclonal antibodies.

Whole cell lysates of Neuro2A  cells were applied to SDS-PAGE. The immunoblots were probed with the indicated monoclonal antibodies for 1h at RT and developed by ECL (exp. time 30 sec).

Whole cell lysates of serum-starved tumor cells (20.000 cells per lane) were applied to SDS-PAGE and transferred to PVDF membrane. Immunoblot were probed with (4) mab Rictor-1G11 (0.5µg/ml) and (5) mab Raptor-10E10 (0.5µg/ml) for 1h at RT.