autologous blood and marrow transplantation - Blog Science ...

98 Chapter 2: ALL
5:01 2.5:1 1.25:1 0.6:1
E:T Ratio
Figure 2. Bcl-2 overexpression inhibits LAK cytotoxicity of697 ALL. LAK cells were gen­
erated by incubation of random donor peripheral blood mononuclear cells at 10 6
lmL in RPMI
with 10% fetal bovine serum (FBS) (CM) with addition of5000 U recombinant human IL-2/mL
for 5 days. The growth inhibition assay is performed by proliferation of target cells both with
and without addition of irradiated effector cells. Targets are placed in round-bottomed 96-well
plates at 25,000 per well in triplicate in CM. Effectors are irradiated (30 Gy) and plated with
targets at four different effector .target ratios in triplicate. Background proliferation from irra­
diated effectors is calculatedfrom parallel wells containing only effectors. Proliferation of tar­
gets only is performed with six replicates. After 4-hour coincubation, all wells are pulsed with
1 mCi [ 3
H]thymidine overnight and cpm obtained by standard liquid scintillation counting.
Percentage growth inhibition is scored after subtraction of effectors alone and is based on cpm
of targets alone. Shown are mean and standard deviation for five separate experiments.
5:01
2.5:1 1.25:1
E:T Ratio
1-2
-697Bcl-2
Figure 3. Bcl-2 overexpression has no effect on CTL cytotoxicity of 697 ALL. CTL were
generated by incubation of random donor peripheral blood mononuclear cells at ItflmL in
RPMI with 10% FBS (CM) with addition of 50 U recombinant human IL-2/mlfor 5 days with
irradiated (60 Gy) 697 or 697Bcl-2 stimulators at a 3:1 responder-to-stimulator ratio. The
growth inhibition assay is performed identically to that described in Fig. 2. Shown are mean
and standard deviation for four separate experiments.
.6:1
-697