Journal: Neurology Research International

Article Title: Subarachnoid Transplant of the Human Neuronal hNT2.19 Serotonergic Cell Line Attenuates Behavioral Hypersensitivity without Affecting Motor Dysfunction after Severe Contusive Spinal Cord Injury

doi: 10.1155/2011/891605

Figure Lengend Snippet: Neural and human markers with differentiation of the hNT2.19 cell line in vitro. The hNT2.19 cell line was treated for two weeks with retinoic acid and mitotic inhibitors and lifted to substrate-coated 8-well plastic TC slides for differentiation and immunohistochemistry for neuron-specific markers. As soon as 4 days in vitro, a variety of neural markers appeared, which remained strong until at least 6 wks of differentiation: TuJ1 (a), hNSE (b), NFL (c), NFM (d), and NFH (e). For comparison, the negative control hNT2.6 cell line was cultured similarly as the hNT2.19 cells and is here stained for TuJ1 (f). Magnification bar = 20 nm, (a–f).

Article Snippet: For immunohistochemistry of sectioned spinal cord tissues, the polyclonal antibody anti-5HT (ab10385; dilution 1/100 (in vivo)) was purchased from Abcam Inc, Cambridge, MA, and the antihuman TuJ1 antibody (Neuron-specific class III beta-tubulin) was purchased from Neuromics, Edina, MN (MO15013; dilution 1/100 (in vivo).

Techniques: In Vitro, Immunohistochemistry, Comparison, Negative Control, Cell Culture, Staining

Journal: Neurology Research International

Article Title: Subarachnoid Transplant of the Human Neuronal hNT2.19 Serotonergic Cell Line Attenuates Behavioral Hypersensitivity without Affecting Motor Dysfunction after Severe Contusive Spinal Cord Injury

doi: 10.1155/2011/891605

Figure Lengend Snippet: Transplant of hNT2.19 and hNT2.6 cell lines in the severe contusive SCI model: TuJ1 and 5HT immunohistochemistry. Rats were injured with severe contusive SCI followed at two weeks by hNT2.6 (a, b) or hNT2.19 (c, d) cell grafts. Sagittal spinal cord sections were examined at 8 wks after SCI for evidence of surviving lumbar subarachnoid hNT2.6 (a, b) or hNT2.19 (c, d) cell line grafts, utilizing TuJ1 (a, c) or 5HT (b, d) immunohistochemistry. The hNT2.19 and control hNT2.6 (10 6 cells/injection), which had been differentiated for two weeks in vitro, were injected into the subarachnoid space two weeks after the SCI. Cell graft sites were colocalized with 5HT (b, d) and the human-specific marker TUJ1 (neuron-specific class III β -tubulin; (a, c)). There are many surviving hNT2.19 (c) and hNT2.6 (a) grafted cells visible on the pial surface, which stain for TuJ1 (arrows) at the end of the experiment, 56 days after SCI and about 6 weeks after cell transplant. Adjacent sections with the same grafted hNT2.19 (d) and hNT2.6 cells (b) are stained for 5HT, but only the hNT2.19 cells (d) are labeled for 5HT (arrows).

Article Snippet: For immunohistochemistry of sectioned spinal cord tissues, the polyclonal antibody anti-5HT (ab10385; dilution 1/100 (in vivo)) was purchased from Abcam Inc, Cambridge, MA, and the antihuman TuJ1 antibody (Neuron-specific class III beta-tubulin) was purchased from Neuromics, Edina, MN (MO15013; dilution 1/100 (in vivo).

Techniques: Immunohistochemistry, Control, Injection, In Vitro, Marker, Staining, Labeling

Journal: bioRxiv

Article Title: OrganoLabeling: Quick and accurate annotation tool for organoid images

doi: 10.1101/2024.04.16.589852

Figure Lengend Snippet: Immunochemical characterization of embryoid bodies and 30 days old brain organoids generated under three different conditions. A) Embryoid bodies stained to demonstrate the presence of three distinct germ layers; with α-SMA (mesodermal marker), Nestin (ectodermal marker), Sox17 (endodermal marker) and DAPI (cell nucleus), B) brain organoid generated under static condition stained with Sox2 and Tuj1, C) brain organoid generated under static condition stained with N-cadherin, D) brain organoid generated using orbital shaker stained with Sox2 and Tuj1, E) brain organoid generated using orbital shaker stained with N-cadherin, F) brain organoid generated using microfluidic chip stained with Sox2 and Tuj1, G) brain organoid generated using microfluidic chip stained with N-cadherin. (Right panels; magnification of dashed zones)

Article Snippet: Subsequently, primary antibodies (α-SMA (Cell Signaling Technology, Cat no.48938S), Nestin (Proteintech, Cat no. 19483-1-AP), Sox17 (Abcam, Cat no. Ab84990), Sox2 (Bioss, Cat no. bs-0523R), Tuj1 (R&D Systems, Cat no. MAB1195), N-cadherin (Cell Signaling Technology, Cat no. 13116T) were added according to related samples and incubated for overnight at 4 °C.

Techniques: Generated, Staining, Marker

Journal: Brain Sciences

Article Title: Neurite Growth and Polarization on Vitronectin Substrate after in Vitro Trauma is not Enhanced after IGF Treatment

doi: 10.3390/brainsci8080151

Figure Lengend Snippet: Example of a grid used for measurements of neurite density in primary hippocampal neuron cultures. A cross covering one or more neurites was considered one observation. A cross placed above a cell nucleus was not counted. To exemplify, arrows mark crosses that should be considered one observation, while arrowheads marks crosses that should be considered no observation. (red = beta III tubulin, green/yellow = MAP2).

Article Snippet: Thereafter, they were incubated in a humid chamber at 4 °C for 24 h with rabbit polyclonal antibody against MAP2 (Millipore, dilution 1:1000) and mouse monoclonal antibody against beta III (anti-beta III tubulin antibody neuronal marker, Abcam, dilution 1:2000).

Techniques:

Journal: Brain Sciences

Article Title: Neurite Growth and Polarization on Vitronectin Substrate after in Vitro Trauma is not Enhanced after IGF Treatment

doi: 10.3390/brainsci8080151

Figure Lengend Snippet: Hippocampal neurons grown on the vitronectin substrate showed an early increase in neurite growth compared with the controls. At 2 and 3 days there was significantly (*) more neurites per area (calculation of neurite density explained in and in the methods section) when neurons were grown on vitronectin compared to the controls ( n = 6 (6 wells per time point and treatment), two-way ANOVA, Bonferroni post hoc test, p < 0.05) (red = beta III tubulin, green/yellow = MAP2).

Article Snippet: Thereafter, they were incubated in a humid chamber at 4 °C for 24 h with rabbit polyclonal antibody against MAP2 (Millipore, dilution 1:1000) and mouse monoclonal antibody against beta III (anti-beta III tubulin antibody neuronal marker, Abcam, dilution 1:2000).

Techniques:

Journal: Brain Sciences

Article Title: Neurite Growth and Polarization on Vitronectin Substrate after in Vitro Trauma is not Enhanced after IGF Treatment

doi: 10.3390/brainsci8080151

Figure Lengend Snippet: The addition of IGF-1/IGFBP-2 to cultured hippocampal neurons did not show any increase in neurite numbers per area, but rather showed a decrease compared with the controls and in total and a marked decrease in neurites per area compared with the hippocampal neurons grown on standard coating or vitronectin ( n = 6 (6 wells per time point and treatment), two-way ANOVA with Bonferroni post hoc test, p < 0.05) (red = beta III Tubulin, green/yellow = MAP2).

Article Snippet: Thereafter, they were incubated in a humid chamber at 4 °C for 24 h with rabbit polyclonal antibody against MAP2 (Millipore, dilution 1:1000) and mouse monoclonal antibody against beta III (anti-beta III tubulin antibody neuronal marker, Abcam, dilution 1:2000).

Techniques: Cell Culture

Journal: Brain Sciences

Article Title: Neurite Growth and Polarization on Vitronectin Substrate after in Vitro Trauma is not Enhanced after IGF Treatment

doi: 10.3390/brainsci8080151

Figure Lengend Snippet: After in vitro trauma to cultured hippocampal neurons, neurite extension was calculated at the border between viable cells and the central area of injury. Neurons regrew neurites slightly better on vitronectin than on the standard coating. At 1, 4, and 7 days after trauma, there were significantly (*) more neurites when neurons were grown on vitronectin compared to the control coating ( n = 6 (6 wells per time point and treatment), two-way ANOVA with Bonferroni post hoc test, p < 0.05) (red = beta III tubulin, green/yellow = MAP2).

Article Snippet: Thereafter, they were incubated in a humid chamber at 4 °C for 24 h with rabbit polyclonal antibody against MAP2 (Millipore, dilution 1:1000) and mouse monoclonal antibody against beta III (anti-beta III tubulin antibody neuronal marker, Abcam, dilution 1:2000).

Techniques: In Vitro, Cell Culture, Control

Journal: Brain Sciences

Article Title: Neurite Growth and Polarization on Vitronectin Substrate after in Vitro Trauma is not Enhanced after IGF Treatment

doi: 10.3390/brainsci8080151

Figure Lengend Snippet: After in vitro trauma and treatment with IGF-1/IGFBP-2, hippocampal neurons neurite extension was calculated at the border between viable cells and the central area of injury. No difference in neurite extensions was shown on vitronectin compared to the standard coating, and a decrease in neurite density was shown compared to neurons cultured without IGF-1/IGFBP-2 treatment ( n = 6 (6 wells per time point and treatment), two-way ANOVA with Bonferroni post hoc test, p < 0.05) (red = beta III tubulin, green/yellow = MAP2).

Article Snippet: Thereafter, they were incubated in a humid chamber at 4 °C for 24 h with rabbit polyclonal antibody against MAP2 (Millipore, dilution 1:1000) and mouse monoclonal antibody against beta III (anti-beta III tubulin antibody neuronal marker, Abcam, dilution 1:2000).

Techniques: In Vitro, Cell Culture

Journal: International journal of molecular medicine

Article Title: The potential of mouse skin-derived precursors to differentiate into mesenchymal and neural lineages and their application to osteogenic induction in vivo.

doi: 10.3892/ijmm.2011.785

Figure Lengend Snippet: Figure 2. Differentiation of mouse SKPs toward peripheral nervous system lineages. Representative images of differentiated SKPs stained with markers for peripheral neurons (A) and Schwann cells (C). Peripheral neurons were double-labeled for TuJ1 with peripherin. Schwann cells were double-labeled for S100β with GFAP. The yellow in the merged images denotes a cluster of cells positive for both markers, with nuclei stained blue (DAPI). Scale bars=50 µm. Real- time PCR for specific marker genes of peripheral neurons (B) and Schwann cells (D) in total RNA isolated from both cells, as compared with SKPs. Data are expressed as the mean ± SD; *P<0.01. RT-PCR for specific marker genes of peripheral neurons (E) and Schwann cells (F). RT-PCR for GAPDH was used as a loading control, and a reaction without input nucleic acid was run in the first lane as a negative control.

Article Snippet: After blocking with PBS containing 1% BSA for 60 min, cells were incubated with primary antibodies to TuJ1 (1:200; R&D Systems, Minneapolis, MN, USA), peripherin (1:50; Santa Cruz Biotechnology, Santa Cruz, CA, USA), glial fibrillary acidic protein (GFAP) (1:500; Chemicon, Temecula, CA, USA), S100β (1:100), smooth muscle actin α (SMAα) (1:100; Sigma-Aldrich), and desmin (1:100; Dako, Glostrup, Denmark) for 1 h at room temperature and then stained with secondary antibodies (FITC-conjugated goat anti-mouse IgG and Cy3-conjugated goat anti-rabbit IgG).

Techniques: Staining, Labeling, Real-time Polymerase Chain Reaction, Marker, Isolation, Reverse Transcription Polymerase Chain Reaction, Control, Negative Control