- 701183 - Invitrogen Antibodies MAPK3 antibody

Submitted references Pathology of Fibrosis in Crohn's Disease-Contribution to Understanding Its Pathogenesis.

Zidar N, Langner C, Jerala M, Boštjančič E, Drobne D, Tomažič A
Frontiers in medicine 2020;7:167

No comments: Submit comment

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of ERK1 in whole cell extracts from U87-MG, MCF-7, HeLa, HepG2, MDA-MB-231, PC12, MDA-MB-453, and Jurkat cells (lanes 1-8 respectively) using an ERK1 recombinant rabbit monoclonal antibody (Product # 701183) at a dilution of 1 µg/mL. Samples were detected using chemiluminescence (ECL). Results show a band at ~42kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of HeLa (Lane 1), H1975 (Lane 2), HCT 116 (Lane 3), HT-29 (Lane 4), A-431 (Lane 5), A549 (Lane 6), MDA-MB-231 (Lane 7), MCF7 (Lane 8), U-87 MG (Lane 9), Raji (Lane 10) and Jurkat (Lane 11). The blots were probed with Anti-ERK1 Recombinant Rabbit Monoclonal Antibody (Product # 701183, 1 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). A 44 kDa band corresponding to ERK1 was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of ERK1 was performed by loading 30 µg of K562 (lane1), A431 (lane2), HEK-293 (lane3), Jurkat (lane4), PC-12 (lane5) and MDA-MB-231 (lane6) cell lysates using Novex®NuPAGE®4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800), and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5 % skim milk for 1 hour at room temperature. ERK1 was detected at ~44 kDa using ERK1 Recombinant Rabbit Monoclonal Antibody (Product # 701183) at 0.5 µg-1 µg/mL in 2.5 % skim milk at 4°C overnight on a rocking platform. Goat anti-Rabbit IgG-HRP Secondary Antibody (Product # G-21234) at 1:5000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of ERK1 in whole cell extracts from U87-MG, MCF-7, HeLa, HepG2, MDA-MB-231, PC12, MDA-MB-453, and Jurkat cells (lanes 1-8 respectively) using an ERK1 recombinant rabbit monoclonal antibody (Product # 701183) at a dilution of 1 µg/mL. Samples were detected using chemiluminescence (ECL). Results show a band at ~42kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of ERK1 was achieved by transfecting MCF7 cells with ERK1 specific validated siRNAs (Silencer® select Product # s11140). Western blot analysis (Fig a) was performed using whole cell lysates from the ERK1 knock down cells (Lane 3), non-specific scrambled siRNA transfected cells (Lane 2) and untransfected cells (Lane 1). The blots were probed with ERK1 Recombinant Rabbit Monoclonal Antibody (Product # 701183, 1 µg/mL) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). Densitometric analysis of this Western blot is shown in histogram (Fig b). Decrease of signal upon siRNA mediated knock down confirms that antibody is specific to ERK1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of ERK1 was performed using 70% confluent log phase A549 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with ERK1 Rabbit monoclonal Antibody (Product # 701183) at 5 µg/mL in 0.1% BSA and incubated overnight at 4 degree Celsius and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing nuclear and cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of ERK1 was done on 70% confluent log phase U2OS cells. The cells were fixed with 4% paraformaldehyde for 15 minutes; permeabilized with 0.25% Triton X-100 for 10 minutes followed by blocking with 5% BSA for 1 hour at room temperature. The cells were incubated with ERK1 Recombinant Rabbit Monoclonal Antibody (Product # 701183) at 2 µg-4 µg in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor® 488 Goat anti-Rabbit IgG Secondary Antibody (Product # A-11008) at a dilution of 1:400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 594 Phalloidin (Product # A12381). Panel d is a merged image showing cytoplasmic localization of ERK1. Panel e shows no primary antibody. The images were captured at 20X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of ERK1 in HeLa cells using an ERK1 recombinant rabbit monoclonal antibody (Product # 701183) followed by detection using an Alexa Fluor 488-conjugated goat anti-rabbit secondary antibody (green) (Image A). Nuclei were stained using DAPI (Image B) and actin stained with Alexa Fluor 594 phalloidin (red) (image C). Image D is a composite image showing cytoplasmic localization of ERK1.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of ERK1 was done on HeLa cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Tritonª X-100 for 20 minutes, and blocked with 5% BSA for 1 hour at room temperature. Cells were labeled with ABfinityª ERK1 Recombinant Rabbit Monoclonal Antibody (701183, red histogram) or with rabbit isotype control (pink histogram) at 2 µg-4 µg/million cells in 2.5% BSA. After incubation at room temperature for 2-3 hours, the cells were labeled with Alexa Fluor¨ 488 Goat Anti-Rabbit Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune¨ Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of ERK1 in Hela cells using an ERK1 recombinant rabbit monoclonal antibody (Product # 701183). Cells were fixed and permeabilized using FIX & PERM (Product # GAS-004) reagent, and detection was performed using an Alexa Fluor 488 goat anti-rabbit IgG (right peak) compared to an isotype control (left peak).

701183

Antibody data