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Image Search Results

Journal: Nature Communications
Article Title: Neuronal differentiation is associated with a redox-regulated increase of copper flow to the secretory pathway
doi: 10.1038/ncomms10640
Figure Lengend Snippet: ( a ) Immunostaining of PAM (green) in a chicken spinal cord (HH stage 20 embryo). Traverse sections of chick embryos were analysed. Differentiated cells in marginal zones were identified by postmitotic marker isl1/2 (red). Scale bar, 50 μm. ( b ) Intensity profile along the line indicated by the arrow in a shows overlap between PAM and Isl1/2 expression. Medial and lateral borders are represented as M and L, respevtively. ( c ) Schematic of differentiation of SH-SY5Y cells by sequential treatments with retinoic acid (DIF RA) and BDNF (DIF BDNF). ( d ) Differentiation is associated with upregulation of genes involved in copper fluxes to mitochondria and, especially, the secretory pathway. The mRNA levels were determined by ΔΔCt analysis using GAPDH as a reference gene and normalized to non-differentiated cells (red line). Each value is presented as mean±s.d. ( n =3). ( e ) Protein levels for ATP7A and Atox1 increase upon differentiation. Equal amount of protein was loaded to each lane. Neuronal differentiation was verified by upregulation of MAP2. ( f ) Cellular copper content in differentiated SH-SY5Y cells (BDNF) is higher than in non-differentiated (ND) cells. Copper amounts were determined by atomic absorption and normalized to protein amounts. Data from three independent measurements. ( g , h ) ATP7A is localized within TGN and vesicular structures which are distinct from endosomal or lysosomal compartments. Coimmunostaining with TGN was shown as a representative image (see for other images). Nucleus was stained with DAPI (blue). Scale bar, 10 μm. Colocalization of ATP7A and various markers were evaluated using Pearson's product-moment coefficients. Three replicate samples were prepared and analysed. Each value is presented as mean±s.d. ( n =3).
Article Snippet: Primary antibodies used in cell staining were mouse monoclonal anti-FLAG M2 clone (Sigma, F1804),
Techniques: Immunostaining, Marker, Expressing, Staining