Alexa Fluor® 488 Anti-NeuN 抗体 [EPR12763] - Neuronal Marker

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Alexa Fluor® 488 Anti-NeuN antibody [EPR12763] - Neuronal Marker (AB190195)

Image A : composite multiplex immunohistochemistry staining using conjugates panel composed of anti-NeuN (Alexa Fluor^® 488) (ab190195, 1/100 dilution, 5 µg/mL) (green), anti-GFAP (Alexa Fluor^® 555) (ab201735, 1/100 dilution, 5 µg/mL) (magenta) and anti-Synaptophysin (Alexa Fluor^® 647) (ab196166, 1/100 dilution, 5 µg/mL) (white) on Formalin/PFA-fixed paraffin-embedded sections of rat cerebellum. Nuclear DNA was labelled with DAPI (blue).

Image B : Formalin/PFA-fixed paraffin-embedded section of rat cerebellum stained with anti-NeuN (Alexa Fluor^® 488) (ab190195, 1/100 dilution, 5 µg/mL).

Image C : Formalin/PFA-fixed paraffin-embedded section of rat cerebellum stained with anti-GFAP (Alexa Fluor^® 555) (ab201735, 1/100 dilution, 5 µg/mL).

Image D : Formalin/PFA-fixed paraffin-embedded section of rat cerebellum stained with anti-Synaptophysin (Alexa Fluor^® 647) (ab196166, 1/100 dilution, 5 µg/mL).

Immunohistochemistry staining was performed on a Leica Biosystems BOND^® RX instrument. Heat mediated antigen retrieval with EDTA (pH 9.0, 110°C) for 40 mins. The section was then incubated at room temperature for 1 hour and mounted using Dako Fluorescence Mounting Medium^®.

Image acquisition was performed with Leica SP8 confocal microscope. For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Alexa Fluor® 488 Anti-NeuN antibody [EPR12763] - Neuronal Marker (AB190195)

ab190195 staining NeuN in U87-MG cells. The cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab190195 at 1/50 dilution (shown in green) and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin) at 1μg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an Alexa Fluor^® 594 Goat anti-Mouse secondary (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Alexa Fluor® 488 Anti-NeuN antibody [EPR12763] - Neuronal Marker (AB190195)

Overlay histogram showing U-87MG cells stained with ab190195 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab190195, 1/500 dilution) for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) Alexa Fluor® 488 used at the same concentration and conditions as the primary antibody. Unlabelled sample (blue line) was also used as a control.

Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

This antibody gave a positive signal in U-87MG fixed with 4% formaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Alexa Fluor® 488 Anti-NeuN antibody [EPR12763] - Neuronal Marker (AB190195)

ab190195 staining NeuN in NGF-differentiated PC12 cells (7 days). The cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab190195 at 1/50 dilution (shown in green) and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin) at 1μg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an Alexa Fluor^® 594 Goat anti-Mouse secondary (ab150120) at 2 μg/ml (shown in red). Nuclear DNA was labelled in blue with DAPI.

• IHC-Fr

Lab

Immunohistochemistry (Frozen sections) - Alexa Fluor® 488 Anti-NeuN antibody [EPR12763] - Neuronal Marker (AB190195)

IHC image of ab190195 staining in acetone fixed frozen tissue section of normal rat brain.

Non-specific protein-protein interactions were blocked using TBS containing 0.025% (v/v) Triton X-100, 0.3M (w/v) glycine and 3% (w/v) BSA for 1h at room temperature. The section was then incubated with ab190195 (1/50) in TBS containing 0.025% (v/v) Triton X-100 and 3% (w/v) BSA overnight at +4°C. The section was then counterstained and mounted with SlowFade^® Gold Antifade Mountant with DAPI.

The DAPI only control (no antibody) inset shows no autofluorescence, demonstrating that any Alexa Fluor^® 488 signal is dervied directly from bound ab190195. The separate images of ab190195 and DAPI alone, combined with the merged version of both signals, shows predominant co-localisation of the Alexa Fluor^® 488 signal in the nuclei of the hippocampal granular layer.

For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Alexa Fluor® 488 Anti-NeuN antibody [EPR12763] - Neuronal Marker (AB190195)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse hippocampus (fresh frozen) tissue labeling SAP102 with ab325113 at 1/50 (10.0 ug/ml) dilution (White).

Panel A : merged staining of anti-DLG3 (ab325113, white), anti-NeuN (ab190195, green) and anti-GFAP (ab201732, magenta) on mouse hippocampus.
Panel B : anti-DLG3 stained on mouse hippocampus.
Panel C : anti-NeuN stained in neurons of mouse hippocampus.
Panel D : anti-GFAP stained in astrocytes of mouse hippocampus.
The section was incubated in two rounds of staining : in the order of ab325113 and ab190195, ab201732 for 1 hr at room temperature. The nuclear counterstain was DAPI (Blue). The section was then mounted using Fluoromount®. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).