In Situ Cell Death Detection Kit, Fluorescein

Protocol tips

Upstream tips	Protocol tips	Downstream tips
- 2.5 × 10^4 cells per well in 24 well plate were seeded.	- Cells were stained with anti-TDP-43 (1:150), anti-GFP (1:1000), Alexa 647 anti-rabbit (1:1000) and Alexa 488 anti-chicken (1:1000). - The enzyme solution (terminal transferase) and label solution were mixed in a volume ratio of 1:9

Upstream tips
- 2.5 × 10^4 cells per well in 24 well plate were seeded.
Protocol tips
- Cells were stained with anti-TDP-43 (1:150), anti-GFP (1:1000), Alexa 647 anti-rabbit (1:1000) and Alexa 488 anti-chicken (1:1000). - The enzyme solution (terminal transferase) and label solution were mixed in a volume ratio of 1:9

Publication protocol

Deoxynucleotidyl-transferase-mediated dUTP nick end-labeling (TUNEL) staining was performed using an in situ cell death detection kit (Roche Applied Science, IN). Cells were seeded on poly-D-lysine coated cover slips at a density of 2.5 × 104 cells per well in 24 well plate. After treatment, cells were fixed and stained with anti-TDP-43 (1:150; ProteinTech, Chicago, IL), anti-GFP (1:1000; Invitrogen, Carlsbad, CA), Alexa 647 anti-rabbit (1:1000; Invitrogen, Carlsbad, CA) and Alexa 488 anti-chicken (1:1000; Invitrogen, Carlsbad, CA). The enzyme solution (terminal transferase) and label solution were mixed in a volume ratio of 1:9 to obtain the TUNEL reaction mixture. The cells were incubated with the 50 μl TUNEL reaction mixture at 37°C for 1 hour and then washed three times with PBS. Then the cover slips were mounted on glass slides with antifade reagent (Invitrogen, Carlsbad, CA). Images were obtained with an Olympus Fluoview FV1000 Confocal scanning microscope.

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