Veenstra et al. (1998) Cell Death Diff. 5:774-784
This protocol uses the Cell death detection ELISA reagents from Roche / Boehringer Mannheim. Submitted by Gert Veenstra.
Wash embryos 1 x in 25% MMR
Remove excess buffer
Add 10 volumes "incubation buffer", i.e. 50 µl for 5 embryos
Lyse the embryos by gently pipetting up and down using a large blue tip
Use a negative control, as shearing of genomic DNA may give false positive signals
Dilute 10 µl lysate with 190 µl "incubation buffer"
Consider testing other dilutions
Incubate this sample for 30 min. at 4°C
Centrifuge at 13,000 rpm (eppendorf) for 10 min. at 4°C
Remove 160 µl supernatant / embryo, avoiding pellet contamination of the supernatant
Samples can be stored, if not used the same day, in aliquots at -20°C
Do ELISA according to manufacturer's instructions.