Journal: Metabolic Brain Disease

Article Title: Spatial metabolomics reveals the effects of Acanthopanax senticosus on region-specific alterations in neurotransmitters and metabolites levels in the brains of α-syn transgenic Parkinson’s disease model mice

doi: 10.1007/s11011-025-01673-z

Figure Lengend Snippet: Representative images of histopathological and immunohistochemical staining ( n = 3). ( A ) HE staining was used to assess changes in neurons in the substantia nigra of mice (×400), and ( B ) TH + neurons (×200) and ( C ) α-syn neurons (×200) were detected using immunohistochemical staining. ( D ) Analysis of the α-syn areal density in the striatum. ( E ) Positive rate of TH + neurons in the substantia nigra. Compared with the control group, ## P < 0.01, # P < 0.05; Compared with the model group, * P < 0.05

Article Snippet: The tissue sections were evenly covered dropwise with 3% BSA and blocked at room temperature for 30 min. After washing with gentle shaking to remove the blocking solution, primary antibodies against TH (5206 S, CST) and α-syn (2642 S, CST) diluted at a 1:50 ratio were added, ensuring full contact with the tissue sections.

Techniques: Immunohistochemical staining, Staining, Control

Journal: Frontiers in Neurology

Article Title: In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons

doi: 10.3389/fneur.2018.00180

Figure Lengend Snippet: Representative images of immunofluorescence staining of non-tg cortical primary neurons (12 DIV) using antibodies against alpha-synuclein (αSyn) [mAb1338, red, (A) ], VAMP-2 [(green), (B) ], SNAP-25 [(green), (C) ], and syntaxin-1 [(green), (D) ]. DAPI in blue. Scale bar 50 µm.

Article Snippet: The following antibodies were used for the immunofluorescence and PLA experiments, with the same concentration for both techniques: mouse monoclonal mAb1338 recognizing both endogenous mouse αSyn (m-αSyn) and human αSyn (h-αSyn) at 4 μg/ml (R&D Systems, Minneapolis, MN, USA), mouse monoclonal Syn 204 against h-αSyn at 4 μg/ml (Santa Cruz Biotechnology), rabbit monoclonal EPR12790 against VAMP-2 at 2 μg/ml (Abcam, Cambridge, UK), rabbit monoclonal EP3274 against SNAP-25 at 4 μg/ml (Abcam), rabbit polyclonal against syntaxin-1 at 1:1,000 (ABR-Affinity Bioreagents, Golden, CO, USA), rabbit polyclonal AB5622 against microtubule associate protein MAP2 at 1:200 (Merck Millipore, Burlington, MA, USA), and rabbit monoclonal C39A9 against nucleoporin NUP98 at 1:50 (Cell Signaling Technologies, Danvers, MA, USA).

Techniques: Immunofluorescence, Staining

Journal: Frontiers in Neurology

Article Title: In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons

doi: 10.3389/fneur.2018.00180

Figure Lengend Snippet: Characterization of primary neurons from tg A30P mice. Levels of h-alpha-synuclein (αSyn) in lysates of non-tg and A30P neuronal cultures were measured by ELISA, using the h-αSyn specific antibody 4B12 as a capture antibody (A) . Levels of total (t)-αSyn (m-αSyn + h-αSyn) in lysates of non-tg and A30P neuronal cultures were measured by ELISA, using clone 42 as a capture antibody. Two-tailed Student’s t -test, non-significant (ns), error bars represent the SEM (B) . Quantification of h-αSyn positive non-tg and A30P neurons, observed by immunofluorescence with h-αSyn specific antibody Syn 204 (C) . Representative images of immunofluorescence of tg A30P neurons with t-αSyn mAb1338 (D) . Representative images of immunofluorescence with h-αSyn specific antibody Syn 204 of A30P cortical neurons (red). MAP2 neuronal marker (green) and DAPI (blue) (E) . Scale bar 50 µm.

Article Snippet: The following antibodies were used for the immunofluorescence and PLA experiments, with the same concentration for both techniques: mouse monoclonal mAb1338 recognizing both endogenous mouse αSyn (m-αSyn) and human αSyn (h-αSyn) at 4 μg/ml (R&D Systems, Minneapolis, MN, USA), mouse monoclonal Syn 204 against h-αSyn at 4 μg/ml (Santa Cruz Biotechnology), rabbit monoclonal EPR12790 against VAMP-2 at 2 μg/ml (Abcam, Cambridge, UK), rabbit monoclonal EP3274 against SNAP-25 at 4 μg/ml (Abcam), rabbit polyclonal against syntaxin-1 at 1:1,000 (ABR-Affinity Bioreagents, Golden, CO, USA), rabbit polyclonal AB5622 against microtubule associate protein MAP2 at 1:200 (Merck Millipore, Burlington, MA, USA), and rabbit monoclonal C39A9 against nucleoporin NUP98 at 1:50 (Cell Signaling Technologies, Danvers, MA, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Two Tailed Test, Immunofluorescence, Marker

Journal: Frontiers in Neurology

Article Title: In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons

doi: 10.3389/fneur.2018.00180

Figure Lengend Snippet: Representative confocal images of in situ proximity ligation assay (PLA) between alpha-synuclein (αSyn) (mAb1338) and VAMP-2 (red) in non-tg (A,B) and A30P (C,D) cortical primary neurons. Maximum intensity projections of a confocal z-stack including a whole cell were performed to observe the maximum amount of PLA puncta (A,C) . Cross section of the processes allows visualization of the PLA puncta in an orthogonal view [ (B,D) , inlet], scale bar 20 µm. F-actin stained by phalloidin-FITC (green) and DAPI (blue). Quantification of amount of PLA puncta per cell in the soma and processes. One-way ANOVA followed by Bonferroni post hoc test, non-significant (ns), *** P < 0.001. Error bars represent the SEM (E) . Negative control PLA with VAMP-2 antibody only and αSyn antibody (mAb1338) only (F) . Scale bar 20 µm.

Article Snippet: The following antibodies were used for the immunofluorescence and PLA experiments, with the same concentration for both techniques: mouse monoclonal mAb1338 recognizing both endogenous mouse αSyn (m-αSyn) and human αSyn (h-αSyn) at 4 μg/ml (R&D Systems, Minneapolis, MN, USA), mouse monoclonal Syn 204 against h-αSyn at 4 μg/ml (Santa Cruz Biotechnology), rabbit monoclonal EPR12790 against VAMP-2 at 2 μg/ml (Abcam, Cambridge, UK), rabbit monoclonal EP3274 against SNAP-25 at 4 μg/ml (Abcam), rabbit polyclonal against syntaxin-1 at 1:1,000 (ABR-Affinity Bioreagents, Golden, CO, USA), rabbit polyclonal AB5622 against microtubule associate protein MAP2 at 1:200 (Merck Millipore, Burlington, MA, USA), and rabbit monoclonal C39A9 against nucleoporin NUP98 at 1:50 (Cell Signaling Technologies, Danvers, MA, USA).

Techniques: In Situ, Proximity Ligation Assay, Staining, Negative Control

Journal: Frontiers in Neurology

Article Title: In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons

doi: 10.3389/fneur.2018.00180

Figure Lengend Snippet: Representative confocal images of in situ proximity ligation assay (PLA) between alpha-synuclein (αSyn) (mAb1338) and SNAP-25 (red) in non-tg (A,B) and A30P (C,D) cortical primary neurons. Maximum intensity projections of a confocal z-stack including a whole cell were performed to observe the maximum amount of PLA puncta (A,C) . Cross section of the processes allows visualization of the PLA puncta in an orthogonal view [ (B,D) , inlet], scale bar 20 µm. F-actin stained by phalloidin-FITC (green) and DAPI (blue). Quantification of amount of PLA puncta per cell in the soma and processes. One-way ANOVA followed by Bonferroni post hoc test, non-significant (ns), *** P < 0.001. Error bars represent the SEM (E) . Negative control PLA with SNAP-25 antibody only (F) . Scale bar 20 µm.

Article Snippet: The following antibodies were used for the immunofluorescence and PLA experiments, with the same concentration for both techniques: mouse monoclonal mAb1338 recognizing both endogenous mouse αSyn (m-αSyn) and human αSyn (h-αSyn) at 4 μg/ml (R&D Systems, Minneapolis, MN, USA), mouse monoclonal Syn 204 against h-αSyn at 4 μg/ml (Santa Cruz Biotechnology), rabbit monoclonal EPR12790 against VAMP-2 at 2 μg/ml (Abcam, Cambridge, UK), rabbit monoclonal EP3274 against SNAP-25 at 4 μg/ml (Abcam), rabbit polyclonal against syntaxin-1 at 1:1,000 (ABR-Affinity Bioreagents, Golden, CO, USA), rabbit polyclonal AB5622 against microtubule associate protein MAP2 at 1:200 (Merck Millipore, Burlington, MA, USA), and rabbit monoclonal C39A9 against nucleoporin NUP98 at 1:50 (Cell Signaling Technologies, Danvers, MA, USA).

Techniques: In Situ, Proximity Ligation Assay, Staining, Negative Control

Journal: Frontiers in Neurology

Article Title: In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons

doi: 10.3389/fneur.2018.00180

Figure Lengend Snippet: Representative confocal images of in situ proximity ligation assay (PLA) between alpha-synuclein (αSyn) (mAb1338) and syntaxin-1 (red) in non-tg (A,B) and A30P (C,D) cortical primary neurons. Maximum intensity projections of a confocal z-stack including a whole cell were performed to observe the maximum amount of PLA puncta (A,C) . Cross section of the processes allows visualization of the PLA puncta in an orthogonal view ( B,D , inlet), scale bar 20 µm. F-actin stained by phalloidin-FITC (green) and DAPI (blue). Quantification of amount of PLA puncta per cell in the soma and processes. One-way ANOVA followed by Bonferroni post hoc test, non-significant (ns), *** P < 0.001. Error bars represent the SEM (E) Negative control PLA with syntaxin-1 antibody only (F) . Scale bar 50 µm.

Article Snippet: The following antibodies were used for the immunofluorescence and PLA experiments, with the same concentration for both techniques: mouse monoclonal mAb1338 recognizing both endogenous mouse αSyn (m-αSyn) and human αSyn (h-αSyn) at 4 μg/ml (R&D Systems, Minneapolis, MN, USA), mouse monoclonal Syn 204 against h-αSyn at 4 μg/ml (Santa Cruz Biotechnology), rabbit monoclonal EPR12790 against VAMP-2 at 2 μg/ml (Abcam, Cambridge, UK), rabbit monoclonal EP3274 against SNAP-25 at 4 μg/ml (Abcam), rabbit polyclonal against syntaxin-1 at 1:1,000 (ABR-Affinity Bioreagents, Golden, CO, USA), rabbit polyclonal AB5622 against microtubule associate protein MAP2 at 1:200 (Merck Millipore, Burlington, MA, USA), and rabbit monoclonal C39A9 against nucleoporin NUP98 at 1:50 (Cell Signaling Technologies, Danvers, MA, USA).

Techniques: In Situ, Proximity Ligation Assay, Staining, Negative Control

Journal: Frontiers in Neurology

Article Title: In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons

doi: 10.3389/fneur.2018.00180

Figure Lengend Snippet: Representative images of NUP98 immunofluorescence staining (green) in non-tg and A30P alpha-synuclein (αSyn) tg primary neurons, DAPI (blue) (A) . Representative confocal images (maximum intensity projection) of in situ proximity ligation assay (PLA) in non-tg and A30P cortical primary neurons between αSyn (mAb1338) and the NUP98 (red), DAPI (blue) (B) . Scale bar 20 µm.

Article Snippet: The following antibodies were used for the immunofluorescence and PLA experiments, with the same concentration for both techniques: mouse monoclonal mAb1338 recognizing both endogenous mouse αSyn (m-αSyn) and human αSyn (h-αSyn) at 4 μg/ml (R&D Systems, Minneapolis, MN, USA), mouse monoclonal Syn 204 against h-αSyn at 4 μg/ml (Santa Cruz Biotechnology), rabbit monoclonal EPR12790 against VAMP-2 at 2 μg/ml (Abcam, Cambridge, UK), rabbit monoclonal EP3274 against SNAP-25 at 4 μg/ml (Abcam), rabbit polyclonal against syntaxin-1 at 1:1,000 (ABR-Affinity Bioreagents, Golden, CO, USA), rabbit polyclonal AB5622 against microtubule associate protein MAP2 at 1:200 (Merck Millipore, Burlington, MA, USA), and rabbit monoclonal C39A9 against nucleoporin NUP98 at 1:50 (Cell Signaling Technologies, Danvers, MA, USA).

Techniques: Immunofluorescence, Staining, In Situ, Proximity Ligation Assay

Journal: Molecular Neurodegeneration

Article Title: Disrupting α-Synuclein–ClpP interaction restores mitochondrial function and attenuates neuropathology in Parkinson’s disease models

doi: 10.1186/s13024-025-00918-w

Figure Lengend Snippet: CS2 treatment mitigates αSyn PFF induced neurotoxicity. ( A ) Representative images of ClpP, αSyn (Syn204; recognize human αSyn only) and NeuN staining in αSyn PFF-inoculated mouse primary cortical neurons that were pre-treated with TAT or CS2 peptide (scale bar = 30 μm). Quantification of ClpP and αSyn co-localization ( n = 12–13 neurons/group, two-tailed Student’s t test, data are mean ± SEM). ( B - C ) Representative images of ClpP, αSyn (Syn204) and NeuN staining in αSyn PFF-inoculated mouse primary cortical neurons that were pre-treated with TAT or CS2 peptide (scale bar = 30 μm). Neurons were fixed in 4% PFA/4% sucrose/1%Triton-X100 to extract the soluble factions before immunostaining. Quantification of fluorescence intensity of insoluble ClpP ( n > 30 neurons/group, one-way ANOVA with Tukey’s post-hoc test; data are mean ± SEM). ( D - E ) Representative images of β-tubulin (Tuj1) and pS129-αSyn (pS129) staining in αSyn PFF-inoculated mouse primary cortical neurons that were pre-treated with TAT or CS2 peptide (scale bar = 30 μm). Quantification of fluorescence intensity of pS129 ( n = 3–6, one-way ANOVA with Tukey’s post-hoc test; data are mean ± SEM). ( F - H ) Representative images of PSD95, Synapsin1 (SYN1) and pS129 staining in αSyn PFF-inoculated mouse primary cortical neurons that were pre-treated with TAT or CS2 peptide (scale bar = 30 μm). Quantification of PSD95 and SYN1 co-localization ( n > 30 areas were quantitated; one-way ANOVA with Tukey’s post-hoc test; data are mean ± SEM)

Article Snippet: Antibodies against αSyn (Syn204) (2647, 1:500), and MAP2 (4542, 1:1000) were from Cell Signaling Technology.

Techniques: Staining, Two Tailed Test, Immunostaining, Fluorescence