PKC Isoform Antibody Sampler Kit
cat.: HAK21154
Product Type: Antibody Sampler Kit
Species reactivity: Human
Applications: WB
Form: Liquid
Storage condition: Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage buffer: 1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.
Concentration: 1ug/ul
Uniprot #: SwissProt: P17252 Human | P20444 Mouse | P05696 Rat | P05771 Human | P68404 Mouse | P68403 Rat | P05771 Human | P68404 Mouse | P68403 Rat | Q05655 Human | P28867 Mouse | P09215 Rat | Q02156 Human | P16054 Mouse | P09216 Rat | P24723 Human | P23298 Mouse | Q64617 Rat | P05129 Human | P63318 Mouse | P63319 Rat | Q15139 Human | Q62101 Mouse | Q9WTQ1 Rat | Q05513 Human
Images
HAK21154_1.jpg Fig1: Western blot analysis of PKC alpha on different lysates with Rabbit anti-PKC alpha antibody (ET1608-15) at 1/1,000 dilution.

Lane 1: 293T cell lysate (20 µg/Lane)
Lane 2: U-87 MG cell lysate (20 µg/Lane)
Lane 3: NIH/3T3 cell lysate (20 µg/Lane)
Lane 4: C6 cell lysate (20 µg/Lane)
Lane 5: Mouse brain tissue lysate (40 µg/Lane)
Lane 6: Rat brain tissue lysate (40 µg/Lane)

Predicted band size: 77 kDa
Observed band size: 77 kDa

Exposure time: 9 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1608-15) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_2.jpg Fig2: Western blot analysis of PKC alpha on different lysates with Rabbit anti-PKC alpha antibody (ET1608-15) at 1/2,000 dilution.

Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-PKC alpha KD cell lysate

Lysates/proteins at 10 µg/Lane.

Predicted band size: 77 kDa
Observed band size: 77 kDa

Exposure time: 40 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1608-15) at 1/2,000 dilution was used in K1803 at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_3.jpg Fig3: Western blot analysis of PKC beta 1 on different lysates with Rabbit anti-PKC beta 1 antibody (ET1705-30) at 1/5,000 dilution.

Lane 1: L-929 cell lysate (20 µg/Lane)
Lane 2: HeLa cell lysate (20 µg/Lane)
Lane 3: C2C12 cell lysate (20 µg/Lane)
Lane 4: NIH/3T3 cell lysate (20 µg/Lane)
Lane 5: C6 cell lysate (20 µg/Lane)

Predicted band size: 77 kDa
Observed band size: 77 kDa
Exposure time: 2 minutes; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1705-30) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_4.jpg Fig4: Western blot analysis of PKC beta 2 on different lysates with Rabbit anti-PKC beta 2 antibody (ET1609-44) at 1/500 dilution.

Lane 1: K-562 cell lysate (20 µg/Lane)
Lane 2: Raji cell lysate (20 µg/Lane)
Lane 3: Mouse spleen tissue lysate (40 µg/Lane)
Lane 4: Rat spleen tissue lysate (40 µg/Lane)

Predicted band size: 77 kDa
Observed band size: 77 kDa

Exposure time: 25 seconds; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1609-44) at 1/500 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_5.jpg Fig5: Western blot analysis of PKC delta on different lysates with Rabbit anti-PKC delta antibody (ET1701-85) at 1/5,000 dilution.

Lane 1: HeLa cell lysate (15 µg/Lane)
Lane 2: Jurkat cell lysate (15 µg/Lane)
Lane 3: Mouse brain tissue lysate (20 µg/Lane)
Lane 4: Rat brain tissue lysate (20 µg/Lane)

Predicted band size: 78 kDa
Observed band size: 78 kDa

Exposure time: 1 minute 40 seconds; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1701-85) at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_6.jpg Fig6: Western blot analysis of PKC eta on different lysates with Rabbit anti-PKC eta antibody (HA722389) at 1/1,000 dilution.

Lane 1: MCF7 cell lysate (20 µg/Lane)
Lane 2: A431 cell lysate (20 µg/Lane)
Lane 3: COS-1 cell lysate (20 µg/Lane)
Lane 4: Mouse brain tissue lysate (40 µg/Lane)
Lane 5: Rat brain tissue lysate (40 µg/Lane)

Predicted band size: 78 kDa
Observed band size: 78 kDa

Exposure time: 46 seconds; ECL: K1802;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722389) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_7.jpg Fig7: Western blot analysis of PKC gamma on different lysates with Rabbit anti-PKC gamma antibody (ET7108-42) at 1/1,000 dilution.

Lane 1: Mouse brain tissue lysate
Lane 2: Mouse cerebellum tissue lysate
Lane 3: Rat brain tissue lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 78 kDa
Observed band size: 75 kDa

Exposure time: 25 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET7108-42) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_8.jpg Fig8: Western blot analysis of PKC mu on different lysates with Rabbit anti-PKC mu antibody (ET1705-4) at 1/1,000 dilution.

Lane 1: A549-si NT cell lysate
Lane 2: A549-si PKC mu cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 102 kDa
Observed band size: 102 kDa

Exposure time: 40 seconds; ECL: K1802;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1705-4) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_9.jpg Fig9: Western blot analysis of PKC zeta on different lysates with Rabbit anti-PKC zeta antibody (HA722773) at 1/2,000 dilution.

Lane 1: 293T cell lysate
Lane 2: HL-60 cell lysate (negative)
Lane 3: HeLa cell lysate
Lane 4: MDA-MB-231 cell lysate
Lane 5: A431 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 68 kDa
Observed band size: 74 kDa

Exposure time: 1 minute 40 seconds; ECL: K1801;

4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722773) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
HAK21154_10.jpg Fig10: Immunohistochemical analysis of paraffin-embedded rat spleen tissue with Rabbit anti-PKC beta 1 antibody (ET1705-30) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1705-30) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21154_11.jpg Fig11: Immunohistochemical analysis of paraffin-embedded mouse kidney tissue with Rabbit anti-PKC beta 1 antibody (ET1705-30) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1705-30) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21154_12.jpg Fig12: Immunohistochemical analysis of paraffin-embedded mouse spleen tissue with Rabbit anti-PKC beta 2 antibody (ET1609-44) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1609-44) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21154_13.jpg Fig13: Immunohistochemical analysis of paraffin-embedded mouse brain tissue with Rabbit anti-PKC delta antibody (ET1701-85) at 1/200 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1701-85) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
HAK21154_14.jpg Fig14: PKC alpha was immunoprecipitated from 0.2 mg 293T cell lysate with ET1608-15 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using ET1608-15 at 1/1,000 dilution. Anti-Rabbit IgG for IP Nano-secondary antibody (NBI01H) at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: 293T cell lysate (input)
Lane 2: ET1608-15 IP in 293T cell lysate
Lane 3: Rabbit IgG instead of ET1608-15 in 293T cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 2 min; ECL: K1801
HAK21154_15.jpg Fig15: Immunocytochemistry analysis of 293T cells labeling PKC alpha with Rabbit anti-PKC alpha antibody (ET1608-15) at 1/100 dilution.

Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-PKC alpha antibody (ET1608-15) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

Beta tubulin (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.
HAK21154_16.jpg Fig16: Flow cytometric analysis of C6 cells labeling PKC alpha.

Cells were fixed and permeabilized. Then stained with the primary antibody (ET1608-15, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
HAK21154_17.jpg Fig17: Immunohistochemical analysis of paraffin-embedded mouse cerebellum tissue with Rabbit anti-PKC gamma antibody (ET7108-42) at 1/1,000 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET7108-42) at 1/1,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.