nestin pe conjugated antibody  (R&D Systems)

Journal: International Journal of Molecular Sciences

Article Title: High-Contrast Stimulation Potentiates the Neurotrophic Properties of Müller Cells and Suppresses Their Pro-Inflammatory Phenotype

doi: 10.3390/ijms23158615

Figure Lengend Snippet: The combination of high-contrast stimulation and BDNF treatment augmented the neurodifferentiation potential and suppressed the pro-inflammatory phenotype of MCs. Quantification of ( a ) live MCs (fixable viability dye negative cells), ( b ) BrdU incorporation by MCs, ( c ) SOX2 + /nestin + MCs, and ( d ) SOX2 + /GFAP + MCs that were either stimulated by high contrast or unstimulated and cultured in the absence or presence of BDNF at the indicated concentrations (0.1 nM, 1 nM, 10 nM). Densitometric Western blot analysis of p-p65 expression in the ( e ) nuclear and ( f ) cytoplasmic extracts of high-contrast-stimulated and unstimulated MCs that were cultured in the absence or presence of BDNF at the indicated concentrations (0.1 nM, 1 nM, 10 nM). ( g ) Representative Western blot images of p-p65 expression in the nuclear and cytoplasmic extracts of high-contrast-stimulated and unstimulated MCs with or without BDNF treatment. Each experiment was repeated three times. The data are the means ± SEM. The results are presented as fold changes from the control group (unstimulated MCs without BDNF treatment). Differences between the stimulated and unstimulated groups treated with the same concentration of BDNF: + p < 0.05, ++ p < 0.01, +++ p < 0.001. Differences between the high-contrast-stimulated group treated with BDNF and the high-contrast-stimulated group without BDNF treatment: ** p < 0.01, *** p < 0.001. Differences between the unstimulated group treated with BDNF and the unstimulated group without BDNF treatment: ## p < 0.01, ### p < 0.001 (two-way ANOVA followed by Tukey’s post-hoc test).

Article Snippet: Next, cells were fixed using Fixation/Permeabilization Diluent (00-5223-56; Thermo Fisher Scientific) and intracellularly stained with anti-BrdU FITC-conjugated antibody (11-5071-42; clone BU20A, Thermo Fisher Scientific), anti-GFAP Alexa Fluor 647-conjugated antibody (51-9792-82; clone 2.2B10; Thermo Fisher Scientific), anti-nestin PE-conjugated antibody (MA5-23574; clone: 307501; Thermo Fisher Scientific) and anti-SOX2 Alexa Fluor 405-conjugated antibody (IC2018V, clone: 245610, R&D Systems, Minneapolis, MN, USA) according to the manufacturer’s instructions.

Techniques: BrdU Incorporation Assay, Cell Culture, Western Blot, Expressing, Concentration Assay

Journal: iScience

Article Title: Enhanced differentiation of neural progenitor cells in Alzheimer’s disease into vulnerable immature neurons

doi: 10.1016/j.isci.2025.112446

Figure Lengend Snippet: Enhanced neuronal activity and early differentiation in APPswe cerebral organoids (A–C) Quantification of the perimeter (B) and area (C) of control and APPswe cerebral organoids cultured for one month. n = 80–104, from 3 independent differentiations. Scale bar, 500 μm. (D–F) Neuronal activity was measured by a multi-electrode array in control and APPswe cerebral organoids cultured for one month. n = 11–12, from 3 independent differentiations. (G–I) Immunohistochemistry was performed to analyze the distribution of (H) nesitn- and (I) DCX-positive cells in control and APPswe cerebral organoids cultured for one month. n = 9, from 3 independent differentiations. Scale bar, 20 μm. (J–M) Immunoblotting to determine expression levels of SOX2 (K), DCX (L), and TUJ1 (M) in control and APPswe cerebral organoids cultured for one month. n = 10–14, from 3 independent differentiations. (N–P) Flow cytometry analysis was performed on control and APPswe cerebral organoids cultured for one month to assess the expression levels of (O) nestin- and (P) DCX-positive populations. n = 3 from 3 independent differentiations. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001 (Student’s t test). Error bar ± SEM.

Article Snippet: After blocking, the cells were incubated for 2 hours at 4°C with primary antibodies: a nestin PE-conjugated antibody (R&D Systems) and an antibody against DCX.

Techniques: Activity Assay, Control, Cell Culture, Immunohistochemistry, Western Blot, Expressing, Flow Cytometry

Journal: iScience

Article Title: Enhanced differentiation of neural progenitor cells in Alzheimer’s disease into vulnerable immature neurons

doi: 10.1016/j.isci.2025.112446

Figure Lengend Snippet: Increased mitochondrial ROS and its impact on early differentiation in APPswe NPCs (A–C) Immunocytochemistry analysis of nestin (B) and DCX (C) in control and APPswe NPCs and neurons, differentiated for one week. Scale bar, 40 μm. NPCs, n = 30, from 3 independent differentiations; neuron, n = 75, from 6 independent differentiations. (D) Increased MitoSOX signal in APPswe NPCs. n = 3, from 3 independent differentiations. (E) Reduced utilization of OXPHOS in glucose metabolism in APPswe NPCs. n = 5, from 3 independent differentiations. (F–H) Normalization of increased nestin (G) and decreased DCX (H) expression in APPswe NPCs with EUK8 treatment. Scale bar, 40 μm. n = 75, from 5 independent differentiations. (I) Schematic of the experiment categorizing control NPCs into high-ROS and low-ROS groups based on MitoSOX signal intensity. (J–L) No significant differences in nestin and DCX expression between high-ROS and low-ROS NPC groups. Scale bar, 20 μm. n = 40, from 3 independent differentiations. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001 (ANOVA test followed by Dunnett’s post hoc analysis or Student’s t test). Error bar ± SEM.

Article Snippet: After blocking, the cells were incubated for 2 hours at 4°C with primary antibodies: a nestin PE-conjugated antibody (R&D Systems) and an antibody against DCX.

Techniques: Immunocytochemistry, Control, Expressing