Lanes 1-2 & 4-5 : Anti-NMDAR2B antibody [EPR23460-119] (ab254356) at 1/2000 dilution
Lane 3 : Anti-NMDAR2B antibody [EPR23460-119] (ab254356) at 1/1000 dilution

Lane 1 : Mouse hippocampus tissue lysate at 10 µg
Lane 2 : Mouse liver tissue lysate at 20 µg
Lane 3 : Rat brain tissue lysate at 10 µg
Lane 4 : Rat hippocampus tissue lysate at 10 µg
Lane 5 : Rat liver tissue lysate at 20 µg

Secondary
All lanes : Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100000 dilution

Predicted band size: 166 kDa

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

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

Bands of 180KDa and 115KDa respectively represent the full length and endogenously cleaved form.

The expression profile observed is consistent with what has been described in the literature (PMID: 22523092)

Negative control: Liver (PMID: 10049997, 9712644).

Exposure time: 10 seconds.

 

Immunohistochemical analysis of paraffin-embedded human cerebrum tissue labeling NMDAR2B with ab254356 at 1/1000 (0.514 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on the human cerebrum (PMID: 7931566). 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 using ab93684 (Tris/EDTA buffer, pH 9.0).

NMDAR2B was immunoprecipitated from 0.35 mg mouse brain tissue lysate 10 µg with ab254356 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab254356 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: Mouse brain tissue lysate 10 µg.

Lane 2: ab254356 IP in mouse brain tissue lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab254356 in mouse brain tissue lysate

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

Exposure time: 3 minutes.

Sample loaded onto lane 1 was non-boiled as boiling may cause protein aggregates. The expression profile observed is consistent with what has been described in the literature (PMID: 22523092).

Anti-NMDAR2B antibody [EPR23460-119] (ab254356) at 1/1000 dilution + Human hippocampus tissue lysate at 20 µg

Secondary
VeriBlot for IP Detection Reagent (HRP) (ab131366) at 1/100000 dilution

Predicted band size: 166 kDa

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

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

Bands of 180KDa, 150KDa and 115KDa respectively represent the full length and endogenously cleaved form.

The expression profile observed is consistent with what has been described in the literature (PMID: 22523092).

Exposure time: 48 seconds.

Immunohistochemical analysis of paraffin-embedded mouse hippocampus tissue labeling NMDAR2B with ab254356 at 1/1000 (0.514 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on the mouse hippocampus (PMID: 7931566). The section was incubated with ab254356 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.

Immunohistochemical analysis of paraffin-embedded rat hippocampus tissue labeling NMDAR2B with ab254356 at 1/1000 (0.514 ug/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on the rat hippocampus (PMID: 7931566). The section was incubated with ab254356 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.

NMDAR2B was immunoprecipitated from 0.35 mg rat brain tissue lysate 10 µg with ab254356 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab254356 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: Rat brain tissue lysate 10 µg.

Lane 2: ab254356 IP in rat brain tissue lysate.

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab254356 in rat brain tissue lysate.

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

Exposure time: 3 minutes.

Sample loaded onto lane 1 was non-boiled as boiling may cause protein aggregates. The expression profile observed is consistent with what has been described in the literature (PMID: 22523092).