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ImmunocytochemistryAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Immunocytochemistry [1]
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- Product number
- PA5-12936 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Phospho-EGFR (Tyr1016) Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- This antibody is predicted to react with mouse based on sequence homology.
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 400 µL
- Storage
- Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.
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Supportive validation
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- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Phospho-EGFR (Tyr1016) was performed using 70% confluent log phase A-431 cells treated with 200 ng/mL of EGF for 10 minutes. 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 Phospho-EGFR (Tyr1016) Rabbit Polyclonal Antibody (Product # PA5-12936) at 1:250 dilution in 0.1% BSA and incubated overnight at 4 degree 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 membrane localization. Panel e represents cells treated with antagonist, Afatinib (0.5 uM for 6hrs) followed by EGF (200 ng/mL for 10 minutes), showing no Phospho-EGFR staining. Panel f shows untreated cells with no signal. Panel g represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
