| °C in 4-well plates until the initiation of the TUNEL procedure (or proceed to Step 5 with TUNEL procedure). If embryos were stored, wash the embryos again as described in Step 3.
5) Incubate embryos in a 50 µl drop of permeabilization solution [0.5% (v/v) Triton X-100, 0.1% (w/v) sodium citrate] for 30 min at room temperature in a humidified box (a plastic box with wet towels will do). Use of a PAP pen or other hydrophobic pen may aid in forming the permeabilization drop. Incubation in the permeabilization solution too long may cause embryos to lyse.
6) FOR PI: Prepare and label 4 dolphin-nosed tubes. One can add PBS/PVP immediately but don’t add DNase, RNase, propidium iodide or TUNEL mixture until just before before use.
a. DNase (190 µl PBS/PVP 10 µl DNase)
b. RNase A (999 µl PBS/PVP 1 µl RNase)
c. Prepare PI as described above.
d. TUNEL (empty for now)
7) Wash the embryos as described in Step 3. For positive and negative controls go to Step 8. For samples, proceed to Step 9.
上一篇：TUNEL Assay Protocol [Rosen Lab, Baylor College of Medicine] 下一篇：In Situ Apoptosis Detection by TUNEL labeling (Frozen Section) [Wilcox Lab ,Emor
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