Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [3]
- Immunocytochemistry [1]
- Immunohistochemistry [1]
- Flow cytometry [1]
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Validation data
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- Product number
- PA5-24584 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- EGFR Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 400 µL
- Concentration
- 0.5 mg/mL
- 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
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis using an EGFR polyclonal antibody (Product # PA5-24584) in K562 cell lysates (35 µg per lane).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis was performed on membrane enriched extracts (30 µg lysate) of A-431 (Lane 1), H1975 (Lane 2), A549 (Lane 3), HeLa (Lane 4), HCT 116 (Lane 5), HEK-293 (Lane 6), U-87 MG (Lane 7), MDA-MB-231 (Lane 8) and T-47D (Lane 9). The blot was probed with Anti- EGFR Rabbit Polyclonal Antibody (Product # PA5-24584, 1:1000 dilution) 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 180 kDa band corresponding to EGFR was observed in all cell models. 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 using the wet transfer system. 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).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of EGFR (Fig a) was performed by loading 30 µg of A-431 Control (lane1), A-431 EGFR knockout (lane 2) whole cell extracts using Novex® NuPAGE® 10 % Bis-Tris gel, XCell SureLock™ Electrophoresis System (EI0002), Novex® Sharp Pre-Stained Protein Standard (LC5800), and iBlot® Dry Blotting System (IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5% skim milk for 1 hour at room temperature. EGFR was detected at ~ 180 kDa using Anti- EGFR Rabbit Polyclonal Antibody (Product # PA5-24584, 1:1000 dilution) at in 5% skim milk at 4°C overnight on a rocking platform. Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody HRP conjugate (Product # A27036, 1:4000 dilution) was used and chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106). Densitometric analysis of this western blot is shown in histogram (Fig b). Reduction of signal in CRISPR mediated knockout (KO) confirms that antibody is specific to EGFR.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of EGFR was performed using 90% confluent log phase A-431 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 EGFR Mouse monoclonal antibody (Product # PA5-24584) at 5 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) 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 membranous localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry analysis in formalin-fixed, paraffin-embedded human placenta tissue using an EGFR polyclonal antibody (Product # PA5-24584), followed by HRP-conjugated secondary antibody and DAB staining.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Flow cytometry analysis of K562 cells using an EGFR polyclonal antibody (Product # PA5-24584) (right) compared to a negative control cell (left) at a dilution of 1:10-50, followed by a FITC-conjugated goat anti-rabbit antibody