ERK-MAPK Pathway Antibody Kit
cat.: HAK21117
| Product Type: | Antibody Sampler Kit |
|---|---|
| Species reactivity: | Human, Mouse |
| 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. |
| Uniprot #: | SwissProt: Q02750 Human | P31938 Mouse | Q01986 Rat | P27361 Human | P28482 Human | P63085 Mouse | Q63844 Mouse | P21708 Rat | P63086 Rat | Q15418 Human | P18653 Mouse | Q63531 Rat | P05412 Human | P17535 Human | P05627 Mouse | P15066 Mouse | P17325 Rat | P52909 Rat | Q9BUB5 Human | O08605 Mouse |
Images
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Fig1:
Western blot analysis of Phospho-MEK1 (T292) on different lysates with Rabbit anti-Phospho-MEK1 (T292) antibody (ET1612-42) at 1/1,000 dilution. Lane 1: HeLa cell lysate Lane 2: HeLa treated with 100ng/mL Nocodazole for 17 hours cell lysate Lane 3: PC-12 cell lysate Lane 4: PC-12 treated with 100ng/mL Nocodazole for 18 hours cell lysate Lane 5: HeLa treated with 100ng/mL Nocodazole for 17 hours cell lysate, then the membrane treated with λpp for 1 hour Lane 6: PC-12 treated with 100ng/mL Nocodazole for 18 hours cell lysate, then the membrane treated with λpp for 1 hour Lane 7: NIH/3T3 cell lysate Lane 8: NIH/3T3 treated with 100ng/mL Nocodazole for 18 hours cell lysate Lane 9: NIH/3T3 treated with 100ng/mL Nocodazole for 18 hours cell lysate, then the membrane treated with λpp for 1 hour Lysates/proteins at 20 µg/Lane. Predicted band size: 43 kDa Observed band size: 43 kDa Exposure time: 3 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 (ET1612-42) 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. |
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Fig2:
Flow cytometric analysis of NIH/3T3 cells untreated (left) / treated with λpp (right) labeling Phospho-MNK1 (T250 + T255). Cells were fixed and permeabilized. Then stained with the primary antibody (HA723960, 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). |
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Fig3:
Immunocytochemistry analysis of HeLa cells treated with UV for 1 hour labeling Phospho-c-Jun (S73)+JunD (S100) with Rabbit anti-Phospho-c-Jun (S73)+JunD (S100) antibody (HA722800) at 1/500 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 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-Phospho-c-Jun (S73)+JunD (S100) antibody (HA722800) at 1/500 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 (M1305-2, 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. |
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Fig4:
Western blot analysis of Phospho-c-Jun (S73)+JunD (S100) on different lysates with Rabbit anti-Phospho-c-Jun (S73)+JunD (S100) antibody (HA722800) at 1/2,000 dilution. Lane 1: HeLa cell lysate (20 µg/Lane) Lane 2: HeLa treated with 25μg/mL Anisomycin for 30 minutes cell lysate (20 µg/Lane) Lane 3: NIH/3T3 cell lysate (20 µg/Lane) Lane 4: NIH/3T3 treated with 250ng/mL Anisomycin for 30 minutes cell lysate (20 µg/Lane) Lane 5: C6 cell lysate (20 µg/Lane) Lane 6: C6 treated with 25μg/mL Anisomycin for 30 minutes cell lysate (20 µg/Lane) Predicted band size: 36 kDa Observed band size: 36-40 kDa Exposure time: 30 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 (HA722800) 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. |
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Fig5:
Western blot analysis of Phospho-RSK1 (S380) on different lysates with Rabbit anti-Phospho-RSK1 (S380) antibody (ET1610-28) at 1/500 dilution. Lane 1: Hela whole cell lysate Lane 2: Hela (starved overnight) then treated with TPA (200nM 4h) cell lysate Lane 3: NIH/3T3 whole cell lysate Lane 4: NIH/3T3 (starved 24h) then Calyculin A (100nM 15min) cell lysate Lysates/proteins at 10 µg/Lane. Predicted band size: 83 kDa Observed band size: 83 kDa Exposure time: 1 minute; 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 (ET1610-28) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature. |
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Fig6:
Flow cytometric analysis of HeLa cells untreated (left) or treated (right) with PMA for 30 minutes labeling Phospho-Erk1 (T202 + Y204) + Erk2 (T185 + Y187). Cells were fixed and permeabilized. Then stained with the primary antibody (ET1610-13, 1μg/mL) (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). |
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Fig7:
Western blot analysis of Phospho-MNK1 (T250 + T255) on different lysates with Rabbit anti-Phospho-MNK1 (T250 + T255) antibody (HA723960) at 1/5,000 dilution. Lane 1: HeLa cell lysate Lane 2: HeLa treated with 25μg/mL anisomycin for 30 minutes cell lysate Lane 3: HeLa treated with 25μg/mL anisomycin for 30 minutes cell lysate, then the membrane treated with λpp for 1 hour Lane 4: NIH/3T3 starved for 24 hours cell lysate Lane 5: NIH/3T3 starved for 24 hours then treated with 10% FBS for 30 minutes cell lysate Lane 6: NIH/3T3 starved for 24 hours then treated with 10% FBS for 30 minutes cell lysate, then the membrane treated with λpp for 1 hour Lysates/proteins at 20 µg/Lane. Predicted band size: 51 kDa Observed band size: 45 kDa Exposure time: 1 minute 16 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 (HA723960) 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. |
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Fig8:
Immunocytochemistry analysis of NIH/3T3 cells treated with 200nM PMA for 30 minutes labeling Phospho-Erk1 (T202 + Y204) + Erk2 (T185 + Y187) with Rabbit anti-Phospho-Erk1 (T202 + Y204) + Erk2 (T185 + Y187) antibody (ET1610-13) at 1/100 dilution. Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 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-Phospho-Erk1 (T202 + Y204) + Erk2 (T185 + Y187) antibody (ET1610-13) 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 (M1305-2, 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. |
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Fig9:
Western blot analysis of Phospho-Erk1 (T202 + Y204) + Erk2 (T185 + Y187) on different lysates with Rabbit anti-Phospho-Erk1 (T202 + Y204) + Erk2 (T185 + Y187) antibody (ET1610-13) at 1/5,000 dilution and competitor's antibody at 1/5,000 dilution. Lane 1: SH-SY5Y cell lysate Lane 2: SH-SY5Y treated with 100ng/mL hβ-NGF for 10 minutes cell lysate Lane 3: PC-12 cell lysate Lane 4: PC-12 treated with 100ng/mL hβ-NGF for 10 minutes cell lysate Lysates/proteins at 15 µg/Lane. Predicted band size: 41/43 kDa Observed band size: 41/43 kDa Exposure time: 1 minute 50 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 (ET1610-13) at 1/5,000 dilution and competitor's antibody at 1/5,000 dilution were used in 5% BSA at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. |
Note: All products are “FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE”.