All lanes : Anti-Phospholipase C beta 1/PLCB1 + Phospholipase C beta 3/PLCB3 antibody [EPR18714] (ab184743) at 1/5000 dilution

Lane 1 : Human Phospholipase C beta 1 fragment recombinant protein
Lane 2 : Human Phospholipase C beta 3 fragment recombinant protein

Lysates/proteins at 0.01 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 139 kDa


Exposure time: 10 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

Human Phospholipase C beta 1 recombinant protein fragment contains aa518-656 with a His-Tag®. Human Phospholipase C beta 3 recombinant protein fragment contains aa590-706 with a His-Tag®. The immunogen of this product has low homology with Phospholipase C beta 2 and Phospholipase C beta 4.

All lanes : Anti-Phospholipase C beta 1/PLCB1 + Phospholipase C beta 3/PLCB3 antibody [EPR18714] (ab184743) at 1/1000 dilution

Lane 1 : Mouse cerebellum lysate
Lane 2 : Rat cerebellum lysate
Lane 3 : Human fetal brain lysate

Lysates/proteins at 20 µg per lane.

Secondary
Lanes 1-2 : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution
Lane 3 : Goat Anti-Rabbit IgG Peroxidase conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

Predicted band size: 139 kDa
Observed band size: 150 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.
The molecular weight observed is consistent with what has been described in the literature (PMID: 8429045, 17298601). Phospholipase C beta 1 selectively expresses in the brain which has been described in the literature (PMID: 2468162).

All lanes : Anti-Phospholipase C beta 1/PLCB1 + Phospholipase C beta 3/PLCB3 antibody [EPR18714] (ab184743) at 1/1000 dilution

Lane 1 : Mouse brain lysate
Lane 2 : Mouse heart lysate
Lane 3 : Mouse kidney lysate
Lane 4 : Mouse spleen lysate
Lane 5 : Rat brain lysate
Lane 6 : Rat heart lysate
Lane 7 : Rat kidney lysate
Lane 8 : Rat spleen lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

Predicted band size: 139 kDa
Observed band size: 150 kDa why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1,2,3 & 4: 30 seconds; Lane 5,6,7 & 8:10 seconds.

The molecular weight observed is consistent with what has been described in the literature (PMID: 8429045, 17298601). Phospholipase C beta 1 selectively expresses in the brain which has been described in the literature (PMID: 2468162).

All lanes : Anti-Phospholipase C beta 1/PLCB1 + Phospholipase C beta 3/PLCB3 antibody [EPR18714] (ab184743) at 1/1000 dilution

Lane 1 : HCT 116 (Human colorectal carcinoma cell line) whole cell lysate
Lane 2 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
Lane 3 : U-87 MG (Human glioblastoma-astrocytoma epithelial cell line) whole cell lysate
Lane 4 : SH-SY5Y (Human neuroblastoma cell line from bone marrow) whole cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 139 kDa
Observed band size: 150 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.

Immunohistochemical analysis of paraffin-embedded Human cerebrum tissue labeling Phospholipase C beta 1 + Phospholipase C beta 3 with ab184743 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Mainly cytoplasm staining on neurons of the normal Human cerebrum is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Mouse cerebellum tissue labeling Phospholipase C beta 1 + Phospholipase C beta 3 with ab184743 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on purkinje cells of the Mouse cerebellum is observed.

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Rat cerebellum tissue labeling Phospholipase C beta 1 + Phospholipase C beta 3 with ab184743 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Cytoplasm staining on purkinje cell of the Rat cerebellum is observed. 

Counter stained with Hematoxylin.

Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is ab97051 at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Human kidney tissue labeling Phospholipase C beta 1 + Phospholipase C beta 3 with ab184743 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Human kidney. Counter stained with Hematoxylin.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling Phospholipase C beta 1 + Phospholipase C beta 3 with ab184743 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Mouse liver. Counter stained with Hematoxylin.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded Rat testis tissue labeling Phospholipase C beta 1 + Phospholipase C beta 3 with ab184743 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Negative staining on Rat testis. Counter stained with Hematoxylin.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Phospholipase C beta 1 + Phospholipase C beta 3 were immunoprecipitated from 1mg of Mouse brain whole cell lysate with ab184743 at 1/20 dilution.

Western blot was performed from the immunoprecipitate using ab184743 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

Lane 1: Mouse brain whole cell lysate, 10ug (Input).

Lane 2: ab184743 IP in Mouse brain whole cell lysate.

Lane 3: Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) instead of ab184743 in Mouse brain whole cell lysate.

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

Exposure time: 30 seconds.