Anti-NMDAR1 antibody [EPR23397-66]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

Immunohistochemical analysis of paraffin-embedded Human cerebrum tissue labeling NMDAR1 with ab274377 at 1/100 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on human cerebrum (PMID : 27391811). The section was incubated with ab274377 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 solution2) for 20 mins.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling NMDAR1 with ab274377 at 1/100 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Negative control : no staining on human liver (PMID : 10480904). The section was incubated with ab274377 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 solution2) for 20 mins.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling NMDAR1 with ab274377 at 1/100 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on mouse cerebrum (PMID : 27391811). The section was incubated with ab274377 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 solution2) for 20 mins.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling NMDAR1 with ab274377 at 1/100 dilution followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Cytoplasmic staining on rat cerebrum (PMID : 27391811). The section was incubated with ab274377 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 solution2) for 20 mins.

• WB

Lab

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

False colour image of Western blot : Anti-NMDAR1 antibody [EPR23397-66] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab274377 was shown to bind specifically to NMDAR1. A band was observed at 110 kDa in wild-type Neuro-2a cell lysates with no signal observed at this size in GRIN1 knockout cell line ab281960 (knockout cell lysate ab282987). To generate this image, wild-type and GRIN1 knockout Neuro-2a cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (ab274377) at 1/1000 dilution

Lane 1:

Wild-type Neuro-2a cell lysate at 20 µg

Lane 2:

GRIN1 knockout Neuro-2a cell lysate at 20 µg

Lane 3:

SH-SY5Y unboiled cell lysate at 20 µg

Lane 4:

THP-1 unboiled cell lysate at 20 µg

Lane 5:

Human Brain unboiled cell lysate at 20 µg

Lane 6:

Human liver unboiled cell lysate at 20 µg

Lane 7:

Mouse Brain unboiled cell lysate at 20 µg

Lane 8:

Mouse liver unboiled cell lysate at 20 µg

Observed band size: 110 kDa

false

• WB

Lab

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

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

Samples are non-boiled as boiling may cause protein aggregates.

Exposure time : 15 seconds.

All lanes:

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (ab274377) at 1/1000 dilution

Lane 1:

Human brain tissue lysate at 20 µg

Lane 2:

Human heart tissue lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 105 kDa

Observed band size: 120 kDa

false

• WB

Lab

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

Anti-NMDAR1 antibody [EPR23397-66] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab274377 was shown to bind specifically to NMDAR1. A band was observed at 105-125 kDa in wild-type Neuro-2a cell lysates with no signal observed at this size in GRIN1 knockout cell line ab281960 (knockout cell lysate ab282987). To generate this image, wild-type and GRIN1 knockout Neuro-2a cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween®20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (ab274377) at 1/1000 dilution

Lane 2:

Western blot - Mouse GRIN1 (NMDAR1) knockout Neuro-2a cell line (ab281960)

Lane 2:

Western blot - Mouse GRIN1 (NMDAR1) knockout Neuro-2a cell lysate (ab282987)

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Predicted band size: 105 kDa

false

• WB

Lab

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (AB274377)

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

The molecular weight observed is consistent with what has been described in the literature (PMID : 10818139).

Negative control : liver and heart (PMID : 10480904).

Samples are non-boiled as boiling may cause protein aggregates.

Exposure time : 5.5 seconds.

All lanes:

Western blot - Anti-NMDAR1 antibody [EPR23397-66] (ab274377) at 1/1000 dilution

Lane 1:

Mouse brain tissue lysate at 20 µg

Lane 2:

Mouse liver tissue lysate at 20 µg

Lane 3:

Mouse heart tissue lysate at 20 µg

Lane 4:

Rat brain tissue lysate at 20 µg

Lane 5:

Rat liver tissue lysate at 20 µg

Lane 6:

Rat heart tissue lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 105 kDa

Observed band size: 120 kDa

false