Pluronic f 127

The APOPercentage assay and releasing buffer were purchased from Biocolor (Belfast, Northern Ireland, UK). Dihydrorhodamine-123 (DHR 123) and pentylenetetrazol were obtained from Sigma Aldrich (St. Louis, MO, USA), and Caspase-3 (AC-DEVD-AMC) and Caspase-9 (AC-LEHD-AMC) substrates from Enzo (Lausen, Switzerland). Fura 2 (AM) calcium florescent dye was purchased from Calbiochem (Darmstadt, Germany) and Pluronic^® F-127 from Biovision (San Francisco, CA, USA). A mitochondrial stain 5,50, 6,60-tetrachloro-1,10,3,30- tetraethylbenzimidazolyl carbocyanine iodide (JC-1) and Probenecid were purchased from Santa Cruz (Dallas, TX, USA). EGb 761 was provided by AbdIbr Pharmaceuticals.

Staining of worms with tetramethylrhodamine-phalloidin (catalog # P1951, MilliporeSigma, St. Louis, MO) was performed as described (28 (link), 89 (link)). Samples were observed by epifluorescence using a Nikon Eclipse TE2000 inverted microscope (Nikon Instruments, Tokyo, Japan) with a CFI Plan Fluor ELWD 40 × (NA 0.60) objective. Images were captured by a Hamamatsu ORCA Flash 4.0 LT sCMOS camera (Hamamatsu Photonics, Shizuoka, Japan) and processed by NIS-Elements AR V5.02.01 (Nikon Instruments) and Adobe Photoshop CS3.
To measure fluorescence intensity of AIPL-1::GFP, live worms were mounted and immobilized with 25% Pluronic F-127 (catalog # 2730-50G, Biovision, Milpitas, CA) in M9 buffer containing 0.5 mM levamisole and 0.1% tricaine methanesulfonate. Fluorescence images were captured with the same settings for all samples in NIS-Elements using a Nikon Eclipse TE2000 inverted microscope with a CFI Plan Fluor ELWD 40 × (NA 0.60) objective. Fluorescence intensity of each worm was determined using ImageJ as average fluorescence intensity at 10 randomly selected points within the cytoplasm of the body wall muscle in the head region.