Lidocaine ( 10−6to 10−4m) was applied 15 min before addition of phenylephrine.
C60-P or C60(OH)36 was added to responder splenocytes 30 min before addition of the stimulator splenocytes.
ABTS and 0.1 M citrate phosphate buffer were incubated at the optimum temperature of each protein 2 min before addition of enzyme.
GG918 was added to a final concentration of 0, 0.001, and 0.1 μM to the irradiated and not-irradiated cell suspensions at 5 min before addition of 99mTc MIBI.
Graphite oxide (250 mg) was added in 250 mL DI water and stirred for 30 min before addition of 1.4 g NaBH4, and the mixture was kept at 80°C for 1 h.
The cell walls were digested by addition of lysozyme (1 mg/ml) on ice and the samples were then heated at 95°C for 5 min before addition of phenol.
For inhibition experiments, azide-free anti-TRAIL antibody 2E5 (5 µg/ml; Alexis, San Diego, CA) was added to effector cells 45 min before addition of target cells.
Blocking of type I IFN receptor was performed by pre-incubating PBMC with 5 µg/ml anti-IFNAR (Invitrogen) for 30 min before addition of AT-2 HIV.
For supershifting experiments, 1.5 µl of DRIL1 antibody was incubated with the extracts for 30 min before addition of the radiolabeled probe.
The enzyme papain was preactivated in 1.1 mM EDTA, 0.067 mM mercaptoethanol, and 5.5 mM cysteine-HCl for 30 min before addition.
Furthermore, a specific HNF6 and/or HNF4α antibody (Santa Cruz Biotechnology Inc., Heidelberg, Germany) was added to the reaction mix 10 min before addition of the labelled probe.
