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Autophagy Watch

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  1. What can I do to induce starvation?
    In NRK cells, starvation can be induced by changing the media to Hank's Balanced Salt Solution (serum-free) and incubating for 2 – 4 hours. Serum-free DMEM (Dulbecco's modified Eagle's medium) can be used, but the induction is weaker because DMEM contains amino acids.
  2. Tell me more about the inhibitors.
    The well-known anti-malarial drug chloroquine has long been used as an inhibitor of lysosomal activity. Today, its efficacy as an anti-cancer drug is being studied. Bafilomycin A1 is a specific autophagy inhibitor used by many autophagy researchers.
    Another commonly used inhibitor wortmannin (not included in this kit) blocks autophagy at an earlier stage.
  3. Two anti-LC3 antibodies are included. Are they used for different purposes?
    Anti-LC3 antibody, clone 8E10 is conjugated to HRP and suitable for WB. For other applications, such as IC and IP, use clone 4E12.
  4. Can you tell me the details of the experimental protocol for LC3 detection by Western blotting?
    Please refer to the Autophagy (LC3) FAQ.