Anti-PKC delta (phospho S645) antibody [EP1486Y]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Immunohistochemical analysis of paraffin-embedded Human bladder carcinoma labeling PKC delta with ab108972 at 1/500 dilution (0.60 µg/ml). Positive staining on human bladder carcinoma without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab108972 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval was performed with Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Immunohistochemical analysis of paraffin-embedded Human spleen carcinoma labeling PKC delta with ab108972 at 1/500 dilution (0.60 µg/ml). Positive staining on human spleen carcinoma without alkaline phosphatase treatment (image A). No signal was detected when tissues were treated with alkaline phosphatase (image B). The section was incubated with ab108972 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval was performed with Bond™ Epitope Retrieval Solution 2 (pH 9.0) for 20 minutes.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Immunohistochemical analysis of PKC delta in paraffin embedded Human breast carcinoma tissue, using ab108972 at a 1/50 dilution.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Primary ab Dilution 1 : 100,000 dilution, Secondary ab description and code (ab id) Goat Anti-Rabbit IgG, (H+L), HRP conjugated (ab97051), Secondary ab dilution 1 : 20,000 dilution, Blocking buffer and concentration 5% NFDM/TBST, Diluting buffer and concentration 5% NFDM/TBST, Lane 1 : Untreated THP-1 (Human monocytic leukemia cell line) whole cell lysates 15ug, Lane 2 : THP-1 (Human monocytic leukemia cell line) treated with 50nM Calyculin A for 60 minutes whole cell lysates 15ug. Lane 3 : THP-1 (Human monocytic leukemia cell line) treated with 50nM Calyculin A for 60 minutes' whole cell lysates 15ug. Then the membrane was incubated with phosphatase. Lane 4 : None, Observed MW 78 kDa, Exposure time 5 seconds

All lanes:

Western blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (ab108972)

Predicted band size: 78 kDa

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• WB

Lab

Western blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Primary ab Dilution 1 : 100,000 dilution, Secondary ab Goat Anti-Rabbit IgG, (H+L), HRP conjugated (ab97051), Secondary ab dilution 1 : 20,000 dilution,

Blocking buffer and concentration 5% NFDM/TBST, Diluting buffer and concentration 5% NFDM/TBST,

Lane 1 : THP-1 whole cell lysate treated with Calyculin A with no peptides,

Lane 2 : THP-1 whole cell lysate treated with Calyculin A with PKCd S645Pho peptides,

Lane 3 : THP-1 whole cell lysate treated with Calyculin A with PKCd unmodified peptides,

Observed MW 78 kDa,

Exposure time 10 seconds

All lanes:

Western blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (ab108972)

Predicted band size: 78 kDa

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• WB

Lab

Western blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Lanes 1, 5 and 9 : Wild-type HAP1 cell lysate (20 μg)
Lanes 2, 6 and 10 : PKC delta knockout HAP1 cell lysate (20 μg)
Lanes 3, 7 and 11 : A431 cell lysate (20 μg)
Lanes 4, 8 and 12 : HeLa cell lysate (20 μg)
Lanes 1, 2, 3 and 4 : Green signal from target – ab108972 observed at 78 kDa
Lanes 5, 6, 7 and 8 : Red signal from loading control – ab8245 observed at 37 kDa
Lanes 9, 10, 11 and 12 : Merged (red and green) signal

ab108972 was shown to specifically react with PKC delta when PKC delta knockout samples were used. Wild-type and PKC delta knockout samples were subjected to SDS-PAGE. ab108972 and ab8245 (loading control to GAPDH) were diluted 1/10000 and 1/2000 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 1/10000 dilution for 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (ab108972)

Predicted band size: 78 kDa

false

• ELISA

Lab

ELISA - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Antigen PKCd S645Pho peptide, PKCd S645Pho unmodified peptide, Antigen concentration 1000ng/ml, Primary antibody concentration range 0~1000ng/ml, Secondary antibody Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG(H+L), Secondary antibody concentration 1 : 2500 dilution

• Dot

Lab

Dot Blot - Anti-PKC delta (phospho S645) antibody [EP1486Y] (AB108972)

Primary ab dilution 1 : 1000 dilution (2.355 μg /ml), Secondary ab description and code (ab id) Goat Anti-Rabbit IgG, (H+L), HRP conjugated (ab97051), Secondary ab dilution 1 : 100,000 dilution, Blocking buffer and concentration 5% NFDM/TBST, Diluting buffer and concentration 5% NFDM /TBST, Lane 1 : PKCd S645Pho peptides, Lane 2 : PKCd unmodified peptides, Lane 3 : None, Lane 4 : None, Observed MW N/A, Exposure time 3 minutes