|
Download
article
203kb
pdf? |
|
ELISA In Situ: A Sensitive Tool for Detecting
Release of Endogenous BDNF
By Agnieszka Balkowiec,
M.D., Ph.D., and David M. Katz, Ph.D.
Department of Neurosciences, Case Western Reserve University School of Medicine, 10900
Euclid Avenue, Cleveland, OH 44106, USA
To measure release of endogenous BDNF from neurons, we developed an in vitro
model using dissociate cultures of rat primary sensory ganglion cells and a modified
ELISA, termed ELISA in situ (1). Nodose-petrosal ganglion (NPG) neurons were grown in 96
well ELISA plates precoated with Promega Anti-BDNF mAb to capture released BDNF, which we
subsequently detected using the antibody sandwich format of the Promega BDNF Emax®
ImmunoAssay System (Cat.# G7611, G7610). Release was measured in response to either
patterned electrical field stimulation or chronic depolarization with elevated
extracellular potassium and compared with basal release in the absence of depolarizing
stimuli. Incorporation of the coating antibody into the culture system dramatically
increased detectability of BDNF in both control and stimulated cultures. The efficacy of
the in situ assay appears to be related primarily to rapid capture of released
BDNF.
|
