Anti-SNAP25 antibody [RM1207]

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized SNAP25 KO SH-SY5Y (SNAP25 knock out human neuroblastoma epithelial cell) cells labelling SNAP25 with ab320654 at 1/500 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^®488) preadsorbed (ab150081) antibody at 1/1000 dilution (Green).

ab195889 was used to counterstain tubulin at 1/200 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^®488) preadsorbed (ab150081) at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunohistochemical analysis of paraffin-embedded Human cerebrum tissue labeling SNAP25 with ab320654 at 1/5000 (0.102 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human cerebrum. The section was incubated with ab320654 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized SH-SY5Y (human neuroblastoma epithelial cell) cells labelling SNAP25 with ab320654 at 1/500 (1.022 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing membranous staining in SH-SY5Y cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Negative control : HEK-293.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5 ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SNAP25 antibody [RM1207] (AB320654)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized SH-SY5Y (human neuroblastoma epithelial cell) (Right) / HEK-293 (human embryonic kidney epithelial cell) (Left) cells labelling SNAP25 with ab320654 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Negative control : HEK-293.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunohistochemical analysis of paraffin-embedded Human spleen tissue labeling SNAP25 with ab320654 at 1/5000 (0.102 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on human spleen. The section was incubated with ab320654 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SNAP25 antibody [RM1207] (AB320654)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Neuro-2a (mouse neuroblastoma neuroblast) cells labelling SNAP25 with ab320654 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SNAP25 antibody [RM1207] (AB320654)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Mouse primary neuron cells labelling SNAP25 with ab320654 at 1/5000 dilution (0.01ug) / Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized Neuro-2a (mouse neuroblastoma neuroblast) cells labelling SNAP25 with ab320654 at 1/500 (1.022 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing membranous staining in Neuro-2a cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5 ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized PC-12 (rat adrenal gland pheochromocytoma cell) cells labelling SNAP25 with ab320654 at 1/500 (1.022 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing membranous staining in PC-12 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5 ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mouse primary neural/glia cell cells labelling SNAP25 with ab320654 at 1/500 (1.022 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing cytoplasmic staining in mouse primary neural/glia cell (shown in green). The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue).
Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection.
Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).

ab11267 Anti-MAP2 mouse monoclonal antibody was used to counterstain tubulin at 1/500 4ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

-ve control 1 : ab320654 at 1/500 dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) antibody at 1/1000 dilution.
-ve control 2 : ab11267 at 1/500 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunohistochemical analysis of paraffin-embedded Rat spleen tissue labeling SNAP25 with ab320654 at 1/5000 (0.102 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on rat spleen. The section was incubated with ab320654 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labeling SNAP25 with ab320654 at 1/5000 (0.102 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on mouse spleen. The section was incubated with ab320654 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SNAP25 antibody [RM1207] (AB320654)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized PC-12 (rat adrenal gland pheochromocytoma cell) cells labelling SNAP25 with ab320654 at 1/500 dilution (0.1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling SNAP25 with ab320654 at 1/5000 (0.102 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse cerebrum. The section was incubated with ab320654 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling SNAP25 with ab320654 at 1/5000 (0.102 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on rat cerebrum. The section was incubated with ab320654 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument

Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-SNAP25 antibody [RM1207] (AB320654)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized rat primary neural/glia cell cells labelling SNAP25 with ab320654 at 1/500 (1.022 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing cytoplasmic staining in rat primary neural/glia cell (shown in green). The counterstain was observed in magenta. Nuclear DNA was labeled with DAPI (shown in blue).
Confocal scanning Z step was set as 0.3 μm followed by image processing with maximum Z projection.
Image was taken with a confocal microscope(Leica-Microsystems, TCS SP8).

ab11267 Anti-MAP2 mouse monoclonal antibody was used to counterstain tubulin at 1/500 4ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

-ve control 1 : ab320654 at 1/500 dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) antibody at 1/1000 dilution.
-ve control 2 : ab11267 at 1/500 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution.

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-SNAP25 antibody [RM1207] (AB320654)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized Rat primary neuron cells labelling SNAP25 with ab320654 at 1/5000 dilution (0.01ug) / Right compared with a Rabbit monoclonal IgG (ab172730) / Left isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

• WB

Supplier Data

Western blot - Anti-SNAP25 antibody [RM1207] (AB320654)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : heart, kidney and spleen (PMID : 8663154).

The identity of the lower MW band at approximately 20 kDa is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Exposure time : Lanes 1-8 : 6 seconds, Lanes 9-10 : 180 seconds

All lanes:

Western blot - Anti-SNAP25 antibody [RM1207] (ab320654) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 20 µg

Lane 2:

Mouse heart tissue lysate at 20 µg

Lane 3:

Mouse kidney tissue lysate at 20 µg

Lane 4:

Mouse spleen tissue lysate at 20 µg

Lane 5:

Rat brain tissue lysate at 20 µg

Lane 6:

Rat heart tissue lysate at 20 µg

Lane 7:

Rat kidney tissue lysate at 20 µg

Lane 8:

Rat spleen tissue lysate at 20 µg

Lane 9:

Human cerebellum tissue lysate at 20 µg

Lane 10:

Human spleen tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 25 kDa,36 kDa

false

• WB

Lab

Western blot - Anti-SNAP25 antibody [RM1207] (AB320654)

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, ab320654 was shown to bind specifically to SNAP25. Target of interest was observed at 25 kDa in wild-type SH-SY5Y cell lysates (lane 1) with no signal observed at this size in SNAP25 knockout cell line (lane 2) (lane 2, knockout cell line ab280041 / knockout cell lysate ab280100).

All lanes:

Western blot - Anti-SNAP25 antibody [RM1207] (ab320654) at 1/1000 dilution

Lane 1:

Wild-type SH-SY5Y (human neuroblastoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

SNAP25 knockout SH-SY5Y whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 25 kDa

false

Exposure time: 180s