class iii beta tubulin tuj1  (Proteintech)

Journal: Nature cell biology

Article Title: Proteostasis and lysosomal repair deficits in transdifferentiated neurons of Alzheimer's disease.

doi: 10.1038/s41556-025-01623-y

Figure Lengend Snippet: Fig. 1 | Transdifferentiation of human adult fibroblasts into neurons reveals signatures of ageing and AD. a, Human dermal fibroblasts were collected from healthy young and aged, aged/sAD and fAD-PSEN1 donors. Fibroblasts and tNeurons were used for a variety of experiments and the findings were validated in post-mortem brain tissue as well as CSF. b, Levels of DNA damage (left) measured as the number of the nuclear foci of γ-H2AX immunofluorescence (right) in human fibroblasts. White dotted lines outline cell nuclei. Images are representative of three independent experiments; n = 248 young, 304 aged and 252 aged/sAD cells from three donors. Scale bar, 50 μm. c, Age- and AD-related epigenetic alterations. Immunofluorescence of the histone modifications H3K9me3 (left) and H4K16ac (right) in human fibroblasts was measured. Data are from four donors and three independent experiments; H3K9me3, n = 252 young, 241 aged and 237 aged/sAD cells; H4K16ac, n = 157 young, 167 aged and 141 aged/sAD cells. d, Neuronal transdifferentiation efficiency. Representative images of human fibroblasts immunostained for S100A4 and Vimentin, and tNeurons immunostained for Tuj1, GAP43, MAP2 and NeuN with 4,6-diamidino-

Article Snippet: The tNeurons were permeabilized with 0.3% Triton X-100 in PBS for 5 min at room temperature or cold methanol on ice for 20 min and blocked with pre-warmed 5% BSA in PBS at room temperature for 45 min. Next, the tNeurons were incubated overnight in a moisture chamber at 4 °C with the following primary antibodies in 5% BSA: anti-ASC/PYCARD Nature Cell Biology (1:100; Santa Cruz, sc-514414), anti-GAP-43 (1:300; Novus Biologicals, NB300-143), anti-MAP2 (1:1,000; BioLegend, 822501), anti-NeuN (1:500; Abcam, ab177487), anti-NLRP3 (1:200; R&D systems, MAB7578), anti-p62/SQSTM1 (1:500; Proteintech, 18420-1-AP), anti-amyloid-β (1:100; Cell Signaling Technologies, 8243), anti-amyloid-β (1–42) (1:200; Enzo Life Sciences, ADI-905-804-100), anti-APP-CTF (1:200; BioLegend, 802803), anti-HGS (1:200; GeneTex, GTX101718), anti-Hsp27 (HspB1; 1:100; Proteintech, 18284-1-AP), anti-Hsp70 (1:1,000; Abcam, ab45133), anti-LC3B (1:100; Cell Signaling Technologies, 2775), anti-TDP-43 (1:1,000; Proteintech, 12892-1-AP), anti-pTau (AT8) (1:100; Thermo Fisher Scientific, MN1020), anti-pTau (S262) (1:200; FUJIFILM WAKO, 010-27123), anti-pTDP-43 Ser409/410 (1:300; Cosmo Bio, CAC-TIP-PTD-M01 and 1:200; BioLegend, 829901), anti-synapsin-1 (1:200; Abcam, ab64581), anti-tau (1:100; Aves Labs) and anti-β tubulin III (Tuj1) (1:500; Neuromics, CH23005 and 1:1,000; BioLegend, 801201).

Techniques: Immunofluorescence

Journal: ACS Omega

Article Title: LSD Modulates Proteins Involved in Cell Proteostasis, Energy Metabolism and Neuroplasticity in Human Cerebral Organoids

doi: 10.1021/acsomega.4c04712

Figure Lengend Snippet: Characterization of the human cerebral organoids. Nuclei are stained blue with 4′,6-Diamidino-2-Phenylindole, Dihydrochloride (DAPI) in all images. (A) Immunostaining for 5-HT 2A receptors, which colocalize with the neuronal marker MAP2; (B) TUJ1 immunostaining (green), a neuronal marker, accompanied by PAX6 immunostaining in red, which highlights neural progenitors; (C) GFAP immunostaining identifies radial glia and astrocytes. Scale bars represent 400 μm for whole organoid images and 60 μm for zoomed-in images.

Article Snippet: Subsequently, a blocking step was carried out by incubating in a solution containing 3% fetal calf serum in PBS for 1 h. Following blocking, they were subjected to overnight incubation at 4°C with the primary antibody TUJ1 (Neuromics) at a dilution of 1:2000 in the blocking solution.

Techniques: Staining, Immunostaining, Marker

Journal: ACS Omega

Article Title: LSD Modulates Proteins Involved in Cell Proteostasis, Energy Metabolism and Neuroplasticity in Human Cerebral Organoids

doi: 10.1021/acsomega.4c04712

Figure Lengend Snippet: Neurite Outgrowth in Response to LSD: Human brain spheroids were initially plated for 24 h and then subjected to exposure to either 10 or 100 nM LSD for an additional 24 h. For this analysis, the control group included 18 spheroids, while the groups exposed to 10 and 100 nM LSD consisted of 17 and 15 spheroids, respectively. (A) The samples were analyzed using immunofluorescence staining for β-tubulin III (TUJ1; depicted in green) and DAPI staining to highlight nuclei (shown in blue). (B) Sholl analysis was employed to assess neurite outgrowth. An inset in this section provides a schematic of the Sholl analysis method. The analysis was conducted on the mean numbers of crossings for each group of five circles. The quantification was carried out over five independent experiments. Statistical significance is indicated as follows: * p < 0.05, ** p < 0.01, *** p < 0.001, in comparison to the control group. A scale bar indicating 1000 μm is included for reference.

Article Snippet: Subsequently, a blocking step was carried out by incubating in a solution containing 3% fetal calf serum in PBS for 1 h. Following blocking, they were subjected to overnight incubation at 4°C with the primary antibody TUJ1 (Neuromics) at a dilution of 1:2000 in the blocking solution.

Techniques: Control, Immunofluorescence, Staining, Comparison