Biochemical and Biophysical Research Communications
Measurement of dendritic mRNA transport using ribosomal markers
Section snippets
Materials and methods
Dissociated hippocampal neuronal cultures were prepared from post-natal 1 day rat pups as described by Tang et al. [21]. The neurons were plated at a density of 10,000–20,000 cells/cm2 onto a poly-d-lysine-coated coverslip. The cultures were maintained and matured in a growth medium (Neurobasal-A supplemented with B-27 and Glutamax-1, Invitrogen, Carlsbad, CA) for 12 days prior to use. Immunostaining was performed as described by Aakalu et al. [27]. The immunostained cultures were imaged using
Results and discussion
The dendritic transport of RNA granules or mRNAs was measured using the antibodies to ribosome-specific markers, such as ribosomal L4, S6 proteins, and Y10B to immunostain the cultured hippocampal neurons. All the antibodies showed significant immunoreactivity in both the somatic and dendritic regions of the hippocampal neurons (Fig. 1). The immunostaining result in each dendrite shows a diffused pattern, which results from the low magnification in epifluorescent microscopy. GFP-tagged Staufen
Acknowledgment
This work was supported by Grant No. R05-2004-000-10432 from Korean Ministry of Science and Technology.
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