Anti-BACE1 antibody [EPR22802-233]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Immunohistochemical analysis of paraffin-embedded Rat hippocampus tissue labeling BACE1 with ab263901 at 1/4000 dilution (0.19 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on mossy fibers in the hilar region of the dentate gyrus. The section was incubated with ab263901 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 Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Immunohistochemical analysis of paraffin-embedded Mouse hippocampus tissue labeling BACE1 with ab263901 at 1/4000 dilution (0.19 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on mossy fibers in the hilar region of the dentate gyrus. The section was incubated with ab263901 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 Rabbit specific IHC polymer detection kit HRP/DAB (ab209101)..

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded Rat hippocampus tissue staining SYNPR with ab314136 at a 1 : 8000 (0.06 ug/ml) dilution, BACE1 with ab263901 at 1 : 4000 (0.193 ug/ml) dilution and 5HT6 Receptor with ab315380 at a 1 : 1000 (0.504 ug/ml) dilution followed by secondary antibody Opal Polymer HRP Ms + Rb.

Panel A : merged staining of anti-SYNPR (green; Opal™520), anti-BACE1 (magenta;Opal™570) and anti-5HT6 Receptor (yellow;Opal™690) on rat hippocampus.
Panel B : anti-SYNPR staining the mossy fibers in rat hippocampus.
Panel C : anti-BACE1 staining the mossy fibers in rat hippocampus.
Panel D : anti-5HT6 Receptor staining the pyramidal neurons in rat hippocampus.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab314136, ab263901 and ab315380 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal^™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded Mouse hippocampus tissue staining SYNPR with ab314136 at a 1 : 8000 (0.06 ug/ml) dilution, BACE1 with ab263901 at 1 : 4000 (0.193 ug/ml) dilution and 5HT6 Receptor with ab315380 at a 1 : 1000 (0.504 ug/ml) dilution followed by secondary antibody Opal Polymer HRP Ms + Rb.

Panel A : merged staining of anti-SYNPR (green; Opal™520), anti-BACE1 (magenta;Opal™570) and anti-5HT6 Receptor (yellow;Opal™690) on mouse hippocampus.
Panel B : anti-SYNPR staining the mossy fibers in mouse hippocampus.
Panel C : anti-BACE1 staining the mossy fibers in mouse hippocampus.
Panel D : anti-5HT6 Receptor staining the pyramidal neurons in mouse hippocampus.
Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab314136, ab263901 and ab315380 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal^™ 4-color kit. Image acquisition was performed with Leica SP8 confocal microscope.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

• WB

Supplier Data

Western blot - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Blocking and dilution buffer : 5% NFDM/TBST.

ab181602 was used as a loading control at 1/1000000 dilution.

We suggest optimizing experimental protocols (increasing lysate amount, using lower dilution or higher sensitivity ECL substrate) to improve results.

ab263901 could be an alternative for getting stronger signal in testing cell lines.

Lanes 1 - 3:

Western blot - Anti-BACE1 antibody [EPR19523] (<a href='/en-us/products/primary-antibodies/bace1-antibody-epr19523-ab183612'>ab183612</a>) at 1/1000 dilution

Lanes 4 - 6:

Western blot - Anti-BACE1 antibody [EPR22802-233] (ab263901) at 1/1000 dilution

Lanes 1 and 4:

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

Lanes 2 and 5:

Mouse brain tissue lysate at 20 µg

Lanes 3 and 6:

Neuro-2a (Mouse neuroblastoma cell line) 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 1/20000 dilution

Predicted band size: 56 kDa

Observed band size: 68 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure times.

Lanes 1-2 : 26 seconds;
Lanes 3-6 : 10 seconds.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 17425515).

ab263901 was shown to specifically react with BACE1 in wild-type HAP1 cells as signal was lost in BACE1 knockout cells. Wild-type and BACE1 knockout samples were subjected to SDS-PAGE. ab263901 and ab181602 (Rabbit anti-GAPDH loading control) were incubated 1 hour at room temperature at 1/1000 dilution and 1/200,000 dilution respectively. Blots were developed with Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) secondary antibody at 1/100,000 dilution for 1 hour at room temperature before imaging. The blot was developed on a BIO-RAD^® ChemiDoc™ MP instrument using the ECL technique.

All lanes:

Western blot - Anti-BACE1 antibody [EPR22802-233] (ab263901) at 1/1000 dilution

Lane 1:

Wild-type HAP1 whole cell lysate at 20 µg

Lane 2:

BACE1 knockout HAP1 whole cell lysate at 20 µg

Lane 3:

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

Lane 4:

Mouse brain lysate at 20 µg

Lane 5:

Human brain lysate at 20 µg

Lane 6:

Rat hippocampus 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

Predicted band size: 56 kDa

Observed band size: 70 kDa

false

• WB

Unknown

Western blot - Anti-BACE1 antibody [EPR22802-233] (AB263901)

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure time : 3 seconds.

All lanes:

Western blot - Anti-BACE1 antibody [EPR22802-233] (ab263901) at 1/1000 dilution

All lanes:

His-tagged human BACE1 recombinant protein (aa22-460 &aa46-460), 10 ng

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

Predicted band size: 56 kDa

Observed band size: 65-67 kDa,68-70 kDa

false