Anti-IP3 receptor antibody [EPR4537]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IP3 receptor antibody [EPR4537] (AB108517)

Immunohistochemical analysis of paraffin-embedded Human cerebellum tissue labeling IP3 with ab108517 at 1/1000 dilution (0.139 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on human cerebellum. The section was incubated with ab108517 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IP3 receptor antibody [EPR4537] (AB108517)

Immunohistochemical analysis of paraffin-embedded Mouse cerebellum tissue labeling IP3 with ab108517 at 1/1000 dilution (0.139 μg/ml) followed by a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on mouse cerebellum. The section was incubated with ab108517 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20mins.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

• IP

Unknown

Immunoprecipitation - Anti-IP3 receptor antibody [EPR4537] (AB108517)

ab108517 (purified) at 1 : 500 dilution (1.124 μg/ml) immunoprecipitating IP3 receptor in Mouse brain lysate.
Lane 1 (input) : Mouse brain lysate 10μg
Lane 2 (+) : ab108517 & Mouse brain lysate
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab32061 in HeLa whole cell lysate
For western blotting, VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1 : 1000 dilution.
Blocking and diluting buffer : 5% NFDM /TBST.

All lanes:

Immunoprecipitation - Anti-IP3 receptor antibody [EPR4537] (ab108517)

Predicted band size: 314 kDa

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• WB

Lab

Western blot - Anti-IP3 receptor antibody [EPR4537] (AB108517)

Lane 1 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : IP3 receptor knockout HAP1 cell lysate (20 μg)
Lane 3 : Human brain tissue lysate (20 μg)
Lane 4 : SH-SY5Y cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab108517 observed at 270 kDa. Red - loading control, ab18058, observed at 124 kDa.

ab108517 was shown to specifically recognize IP3 receptor in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed when IP3 receptor knockout samples were examined. Wild-type and IP3 receptor knockout samples were subjected to SDS-PAGE. ab108517 and ab18058 (loading control to Vinculin) were diluted 1/1000 and 1/10,000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-IP3 receptor antibody [EPR4537] (ab108517)

Predicted band size: 314 kDa

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• WB

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Western blot - Anti-IP3 receptor antibody [EPR4537] (AB108517)

All lanes:

Western blot - Anti-IP3 receptor antibody [EPR4537] (ab108517) at 1/1000 dilution

Lane 1:

Rat brain lysate at 10 µg

Lane 2:

SH-SY5Y lysate at 10 µg

Lane 3:

Mouse brain lysate at 10 µg

Lane 4:

HeLa brain lysate at 10 µg

Predicted band size: 314 kDa

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• OI-RD Scanning

Unknown

OI-RD Scanning - Anti-IP3 receptor antibody [EPR4537] (AB108517)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.