Immunohistochemistry was performed as previously described [34 (link)], except in the case of dpERK antibody, for which testes were dissected and fixed in 10 mM Tris-HCl, pH 6.8, 180 mM KCl, 50 mM NaF, 10 mM NaVO4 and 10 mM β-glycerophosphate as described in [26 (link)] and then treated as in the case of other antibodies. We used the following primary antibodies: guinea pig anti-Tj (1:3000, gift of D. Godt), rabbit anti-Zfh1 (1:5000, gift of R. Lehmann), rabbit anti-Stat92E (1:1000), rabbit anti-Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (1:200, Cell Signaling #9101), mouse anti-βPS Integrin, mouse anti-Eya, mouse anti-Fas3 (all 1:20, Developmental Studies Hybridoma Bank (DSHB), created by the NICHD of the NIH and maintained at The University of Iowa, Department of Biology, Iowa City, IA 52242), goat anti-Vasa (1:100, Santa Cruz Biotechnology), rabbit anti-GFP (1:500, Life Technologies), chicken anti-GFP (1:500, Aves Labs).
Rabbit anti phospho p44 42 mapk erk1 2 thr202 tyr204
Manufactured by Cell Signaling Technology
Sourced in United States
Rabbit anti-Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) is a primary antibody that recognizes the phosphorylated form of p44/42 MAPK (Erk1/2) at the Thr202 and Tyr204 residues. This antibody is used to detect and quantify the activation of the p44/42 MAPK (Erk1/2) signaling pathway.
Automatically generated - may contain errors
Lab products found in correlation
Rabbit anti p44 42 mapk erk1 2, by Cell Signaling Technology (7 mentions)
Protease and phosphatase inhibitor, by Merck Group (2 mentions)
Rabbit anti akt, by Cell Signaling Technology (2 mentions)
Irdye 800cw donkey anti mouse, by LI COR (2 mentions)
Odyssey clx imaging system, by LI COR (2 mentions)
Phosphatase, by Merck Group (2 mentions)
Mouse anti β actin, by LI COR (2 mentions)
Nupage bis tris sds page, by Thermo Fisher Scientific (2 mentions)
M per, by Thermo Fisher Scientific (2 mentions)
Irdye 680rd goat anti rabbit, by LI COR (2 mentions)
Protease, by Roche (2 mentions)
Mouse anti βps integrin, by Developmental Studies Hybridoma Bank (1 mentions)
Mouse anti eya, by Developmental Studies Hybridoma Bank (1 mentions)
Mouse anti fas3, by Developmental Studies Hybridoma Bank (1 mentions)
Goat anti vasa, by Santa Cruz Biotechnology (1 mentions)
Rabbit anti gfp, by Thermo Fisher Scientific (1 mentions)
Chicken anti gfp, by Aves Labs (1 mentions)
Rabbit anti gapdh antibody, by Abcam (1 mentions)
Pd98059, by Cell Signaling Technology (1 mentions)
Rabbit anti phospho igf 1 receptor β tyr1131, by Cell Signaling Technology (1 mentions)
15 protocols using rabbit anti phospho p44 42 mapk erk1 2 thr202 tyr204
1
Immunohistochemistry Protocol for Drosophila Testes
2
Quantifying MEK-ERK Signaling Pathway
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Mitogen-activated protein kinase kinase (MEK) is upstream of ERK. At 30 min after treatment with the MEK inhibitor PD98059 (10 μM; Cell Signaling Technology, Inc., Danvers, MA, USA), the level of phospho-ERK (p-ERK) was detected by WB.
Samples of 10 μg total protein were separated in SDS-PAGE gel. The proteins were then transferred onto a polyvinylidene fluoride (PVDF) membrane. The PVDF membrane was blocked for 2 h at room temperature and then incubated with primary antibodies [rabbit anti-phospho-p44/42 MAPK (ERK1/2; Thr202/Tyr204; 1:1,000; Cell Signaling Technology, Inc.), rabbit anti-p44/42 MAPK (ERK1/2; 1:1,000; Cell Signaling Technology, Inc.) or rabbit anti-GAPDH antibody (1:5,000; Abcam)] overnight with gentle agitation at 4°C. The following day, the membrane was incubated with secondary antibodies [horseradish peroxidase-conjugated goat anti-rabbit IgG, (1:5,000; CoWin Biotech Co., Ltd., Beijing, China)] for 3 h at room temperature. Protein detection was performed using enhanced chemiluminescence.
Samples of 10 μg total protein were separated in SDS-PAGE gel. The proteins were then transferred onto a polyvinylidene fluoride (PVDF) membrane. The PVDF membrane was blocked for 2 h at room temperature and then incubated with primary antibodies [rabbit anti-phospho-p44/42 MAPK (ERK1/2; Thr202/Tyr204; 1:1,000; Cell Signaling Technology, Inc.), rabbit anti-p44/42 MAPK (ERK1/2; 1:1,000; Cell Signaling Technology, Inc.) or rabbit anti-GAPDH antibody (1:5,000; Abcam)] overnight with gentle agitation at 4°C. The following day, the membrane was incubated with secondary antibodies [horseradish peroxidase-conjugated goat anti-rabbit IgG, (1:5,000; CoWin Biotech Co., Ltd., Beijing, China)] for 3 h at room temperature. Protein detection was performed using enhanced chemiluminescence.
3
Western Blot Analysis of Signaling Pathways
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Cells were lysed in RIPA buffer (Cell Signaling; Danvers, MA, USA) supplemented with protease and phosphatase inhibitors (Sigma-Aldrich). Total protein extracts were run on SDS-PAGE and blotted onto nitrocellulose. Blots were probed overnight. The following antibodies were purchased from Cell Signaling: Rabbit anti-phospho-IGF-1 receptor β (Tyr1131) (#3021, 1:200); rabbit anti-IGF-1 receptor β (#3018, 1:250); rabbit anti-phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (#9101, 1:1000); rabbit anti-p44/42 MAPK (Erk1/2) (#9102, 1:1000); rabbit anti-phospho-Akt XP (Ser473) (#9018, 1:1000); rabbit anti-AKT (#9272, 1:1000); rabbit anti-phospho-EGFR (Tyr1148) (#4404, 1:200); and mouse anti-EGFR (#2239, 1:200). Mouse anti-β-actin was purchased from Sigma-Aldrich (AC-15, 1:15,000). All primary antibodies were diluted in 5% BSA/TBS-T. Goat anti-mouse secondary IRDye 800 antibody (#926-32210, 1:10,000) was purchased from Li-Cor Biosciences (Lincoln, NE, USA). Goat anti-rabbit alexa-fluor 680 secondary antibody (#1027681, 1:10,000) was purchased from Invitrogen (Grand Island, NY, USA). Protein bands were detected using the Odyssey Imaging System (Li-Cor Biosciences, Lincoln, NE, USA).
4
Western Blot Analysis of MAPK Signaling
5
Western Blot Analysis of MAPK Signaling
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Protein lysates were prepared in MPER (ThermoScientific, Waltham, MA, USA) containing protease (Roche, Indianapolis, IN, USA) and phosphatase (Sigma-Aldrich, St. Louis, MO, USA) inhibitors. Lysates were resolved by 4-12% NuPage Bis-Tris SDS-PAGE (Invitrogen) and transferred to nitrocellulose. Antibodies: Rabbit anti-p44/42 MAPK (Erk1/2) (1:1,000, #4695 Cell Signaling Technology, Danvers, MA, USA), Rabbit anti-Phospho-p44/42 MAPK (Erk1/2), Thr202/Tyr204 (1:2,000, #4370, Cell Signaling Technology), Mouse anti-β-actin (1:3,000, #926-42212, LI-COR, Lincoln, NE, USA), IRDye 680RD Goat anti-Rabbit (1:15,000, 925-68071, LI-COR) and IRDye 800CW Donkey anti-Mouse (1:15,000, 92532212, LI-COR). Blots were developed using the LI-COR Odyssey CLx Imaging System.
6
Antibody Characterization for Neural Lineage
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The following antibodies with diluent factors were used in this study: chicken anti-GFP (1:1000; catalog no. A10262, Thermo Fisher Scientific), rabbit anti–phospho-S6 ribosomal protein (S235/S236) (1:500; catalog no. 4858, Cell Signaling Technology), rabbit anti–phospho-S6 ribosomal protein (S240/S244) (1:400; catalog no. 5364, Cell Signaling Technology), mouse anti–βIII-tubulin (1:500; catalog no. 4466, Cell Signaling Technology), rabbit anti–phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (1:10,000; catalog no. 4370, Cell Signaling Technology), rabbit anti-SOX2 (1:100; catalog no. ab92494, Abcam), rabbit anti-DCX (1:400; catalog no. ab18723, Abcam), rabbit anti-calretinin (1:100; catalog no. ab702, Abcam), rabbit anti-Fas (1:100; catalog no. ab82419, Abcam), and rat anti-BrdU (1:100; catalog no. ab6326, Abcam).
7
Immunohistochemical Analysis of Lung Tissue
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Paraffin-embedded lung sections (4-µm thickness) were used for this study. Images were taken under polarized light using an upright dry under 40× objective. Tissues were deparaffinized and, after antigen retrieval, permeabilized with 0.4% Triton X-100. Tissues were blocked with 10% BSA and maintained in PBS + 5% BSA + 0.4% Triton X-100 throughout antibody treatments. Primary antibodies were incubated at 4°C overnight, and secondary antibodies were incubated for 1 h at room temperature. Primary antibodies used (1:200 dilution) included rabbit anti-phospho-p44/42 MAPK (Erk1/2; Thr202/Tyr204; #9101S; Cell Signaling Technology); rabbit anti-NF-κB p65 (#8242; Cell Signaling Technology); mouse anti-NFATC2 (#sc-7296, Santa Cruz Biotechnology) mouse anti-CD3 (#sc-7296; Santa Cruz Biotechnology); rabbit anti-Fhl2 (#ab12328; Abcam); and mouse anti-PAR2 (SAM11, #817201; BioLegend). Goat anti-rabbit IgG-A594 or IgG-A488 was used as secondary antibody (1:200 dilution at room temperature). DAPI was used for nuclear counterstaining. ProLong Gold antifade reagent was used as mounting media. Mounted slides were examined with a Leica DM6000 B. Slide Book 6 (3i) was used for analysis and capturing images.
8
Versatile Cell Line Assay Reagents
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HeLa, HEK293T, and RAW 264.7 cells were obtained from American Type Culture Collection, and MEF cells were a gift from A. Hoffmann (University of California Los Angeles). Glutathione–agarose beads and Ni–nitrilotriacetic acid (NTA) agarose beads were purchased from BioBharati LifeScience Pvt Ltd. Mouse anti-HA antibody was purchased from BioLegend. Rabbit anti-NEMO, rabbit anti-IKK2, rabbit anti-His, rabbit anti-Ub, rabbit anti-Myc, rabbit anti-γ-actin, and rabbit anti-ERK2 were purchased from BioBharati LifeScience Pvt Ltd. Mouse anti-p84 was purchased from GeneTex. Mouse anti-GST, rabbit anti-p65/RelA, and rabbit anti-IκBα were from Santa Cruz Biotechnology. Rabbit anti-p100/p52 was a gift from N. Rice (National Cancer Institute; Frederick, MD). Rabbit anti–phospho-SAPK (stress-activated protein kinase)/JNK (Thr183/Tyr185), rabbit anti-SAPK/JNK, rabbit anti–phospho-p38 MAPK (Thr180/Tyr182), rabbit anti-p38 MAPK, rabbit anti–phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204), and rabbit anti–phospho-IKKα/β (Ser177/181) were purchased from Cell Signaling. Horseradish peroxidase–conjugated anti-rabbit and antimouse secondary antibodies were from BioBharati LifeScience Pvt Ltd as was mouse TNF-α. LPS was purchased from Sigma. Mouse LTβR was purchased from Abcam.
9
Stress Response Modulators and Signaling Pathways
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Stress inducers and inhibitors: PD173074, staurosporine, anisomycin, etoposide, brefeldin A, thapsigargin, doxorubicin and Fas ligand were from Sigma-Aldrich (MO, USA). Actinomycin D and camptothecin were from Santa Cruz Biotechnology (TX, USA). ABT 737, BTSA-1, BGJ398, ARQ087 were from Selleckchem (TX, USA), Apo2/TRAIL was from Merck (Darmstadt, Germany). The following primary antibodies were used: rabbit anti-PARP1/2 (9542), rabbit anti-BAX (D2E11) (5023), rabbit anti-phospho-p44/42 MAPK (ERK1/2) (Thr202/Tyr204) (9101) from Cell Signaling Technology (MA, USA), mouse anti-gamma tubulin (T6557) from Sigma-Aldrich, goat anti-FGF1 (sc-1884), mouse His-tag (H3) (sc-8036) and mouse anti-p53 DO-1 (sc-126) from Santa Cruz Biotechnology. Rabbit polyclonal antibodies against FGF1 (OPA1) were generated by Davids Biotechnologie GmbH (Regensburg, Germany) by immunizing rabbits with purified FGF1. The following HRP-conjugated secondary antibodies from Jackson Immuno Research Laboratories (PA, USA) were used: peroxidase affinipure goat anti-rabbit IgG (111-035-144) and anti-mouse IgG (115-035-003).
10
Antibody Immunoblotting for Synaptic Signaling
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The primary antibodies were rabbit anti-phospho-GSK-3α/β (Ser21/9; Cell Signaling, #9331), rabbit anti-GSK-3β (Cell Signaling, #9315), rabbit anti-tubulin (Tuba1; GeneTex, GTX124965), rabbit anti-phospho-p44/42 MAPK (Erk1/2; Thr202/Tyr204; Cell Signaling, #4370), mouse anti-p44/42 MAPK (Erk1/2; Cell Signaling, #9107), rabbit anti-post-synaptic density-95 (PSD-95; GeneTex, GTX80682), and rabbit anti-beta-actin (Abcam, ab8227). The secondary antibodies were HRP-conjugated anti-rabbit (Cell Signaling, 7074S) or HRP-conjugated anti-mouse (Bio-Rad). All antibodies were diluted in 3% milk/TBST, except phospho-specific antibodies, which were diluted in 2.5% bovine serum albumin/TBST.
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