2008 PBL Cat_mech_r.qxd:PBL Catalog.mech 11/29/07 1:50 PM Page 23
Results Discussion
The addition of poly I:C in transfection complexes to A549 cells resulted Type I IFNs are expressed at varying levels in many cell types in
in a dose-dependent increase of RLU activity in the iLite
TM
assay response to a wide variety of stimuli. Furthermore, there are many
suggesting that type I interferons were present in the supernatants. To aspects of immune function that do not involve type I IFNs.
confirm the RLU activity was attributable to type I interferons, a Consequently, one cannot predict a priori if a given stimulus will or will
separate aliquot of the cell culture supernatants were mixed with a not produce type I IFN production.
monoclonal antibody that recognizes the type I interferon receptor This present study showed that a receptor neutralizing antibody used
chain 2 and neutralizes signaling. Separately, the type II receptor chain in conjunction with a rapid cell based assay (iLite
TM
kit) can quickly and
1 antibody was added in a similar fashion. quantitatively determine the levels of type I IFNs in treated cell
The anti-IFNAR2 neutralizing antibody showed near complete supernatants. Based on these results, more in-depth studies such as
inhibition of the RLU signal demonstrating that the activity in the cell ELISA and/or CPE assays with individual IFN type neutralizing antibodies
culture supernatants was due to type I interferon activity (Figure below). are warranted to discern specifically which type I IFNs are present.
Effects of increasing antibody on iLite
TM
reporter activity of poly I:C-treated A549
cell culture supernatants.
U)
(RL
t
s
t
U
ni
gh
Li
e
iv
l
a
t
R
e
[Poly I:C] (µg)
Increasing amounts of poly I:C (blue circles) produced increased
Relative Light Units (RLUs). The addition of anti-human anti-γR1
antibody inhibited RLU levels at all poly I:C test concentrations [red
squares]. Non-specific anti-IFN-γ R1 [green triangles] showed results
similar to samples where no antibody was added.
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