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Briefly, segmented labels of brain neuropils (AL, MBc, LH) were registered onto a reference brain image using affine registration followed by elastic warping.
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Conventional region-of-interest (ROI) analysis based on manual labeling of brain regions is time-consuming and the results could be inconsistent within and among investigators.
One of the major limitations of this work is the lack of anatomical-based evaluation with manual labeling of brain regions which serves as the reference standard for comparing with the results from automated anatomical labeling.
This antibody does not label cells that do not express SV2A [8] and produces no labeling of brain homogenates from SV2A knockout mice [18].
For labeling of brain sections, a series of 10 frozen sagittal sections at approximately 300 µm intervals were stained and quantified per brain.
Double labeling of brain sections was performed using the rabbit anti-GPR17 polyclonal antibody (1∶100) generated as previously described [11] in combination with the selected primary antibody, in PBS 0.01 M, 0.1% Triton X-100.
tTA/rtTA-Oligo#1: 5'-TTTAGCTGTTTCTCCAGGCCACATATGATTAGTTCC-3' tTA/rtTA-Oligo#2: 5'-AGCTGATTTTCCAGGGTTTCGTACTGTTTCTCTGTT-3' tTA/rtTA-Oligo#3: 5'-ATAGAATCGGTGGTAGGTGTCTCTCTTTCCTCTTTT-3' To assess for specificity and rule out non-specific labeling of brain sections, each radiolabeled oligonucleotide was hybridized with a 100-fold excess of unlabelled oligonucleotide.
Immunocytochemistry. Immunohistochemical labelling of brain tissue was performed using the ABC method, as previously described (22).
Fluorescent labeling of brain sections was done according to a modified protocol of the original diOlistic labeling (Pavlowsky et al, 2010) described by Gan et al (2000).
Immunofluorescence labeling of brain sections revealed αv integrin protein expression in the neuroepithelium and Nrp1 expression in blood vessels, with co-localization at points of neuroepithelial-blood vessel contacts (Fig. S6D).
Labeling of brain tissues with QDs (Santra et al. 2005) and applications of QDs in the CNS may present inevasible, unprotected QD-induced risks on the toxin-susceptible CNS.
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