Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [1]
- ELISA [1]
- Immunocytochemistry [1]
Submit
Validation data
Reference
Comment
Report error
- Product number
- M806B - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- EGF Monoclonal Antibody (3A8), Biotin
- Antibody type
- Monoclonal
- Antigen
- Recombinant full-length protein
- Description
- The M806B anti-EGF antibody (biotinylated conjugate of clone 3A8) has successfully been paired as the detection antibody in a sandwich ELISA with coating antibody M805 (Clone 1H11). Typical dilutions for sandwich ELISA: Coat = 1-3 µg/mL and Detection = 0.125-0.5 µg/mL.
- Reactivity
- Human
- Host
- Mouse
- Conjugate
- Biotin
- Isotype
- IgG
- Antibody clone number
- 3A8
- Vial size
- 100 µg
- Concentration
- 0.5 mg/mL
- Storage
- -20°C
No comments: Submit comment
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of Human EGF was performed by loading 2 µg of recombinant human EGF (Lane 1) or supernatant from an EGF expression clone transfected in 293T cells onto a 4-20% Tris-HCl polyacrylamide gel. Proteins were transferred to a PVDF membrane and blocked with 5% Milk/TBST for at least 1 hour. Membranes were probed with an EGF monoclonal antibody recognizing Human EGF (Product # M806) at a dilution of 1:5000 overnight at 4°C on a rocking platform. Membranes were washed in TBS-0.1%Tween 20 and probed with a goat anti-rabbit-HRP secondary antibody (Product # 31430) at a dilution of 1:10,000 for at least one hour. Membranes were washed and chemiluminescent detection performed using Super Signal West Dura (Product # 34075).
- Conjugate
- Biotin
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Sandwich ELISA analysis of Human EGF was performed using the Thermo Scientific ELISA Kit (Product # EHEGF) by coating a blank 96-well microtiter plate with 100 µL per well of Human EGF moncolonal antibody (Product # M805) in duplicate at 1, 3, 4, 5, 7, and 9 µg/mL in DPBS (Product # 28374) and incubating for 12-18 hours at 4C. The plate was aspirated and blocked with 300 µL per well of 4% BSA and 5% sucrose in DPBS for 1 hour at room temperature. Human EGF recombinant protein at 50 µL per well was added in duplicate at 500, 250, 125, 62.5, 31.25, 15.625, 7.8125 and 0 pg/mL for 2 hours at room temperature along with 50 µL of Human EGF biotinylated monoclonal antibody (Product # M806B) in all applicable wells at 0.2 µg/mL for 2 hours at room temperature. The plate was washed with ELISA Wash Buffer (Product # N503) and incubated with 100 µL per well of Streptavidin-HRP (Product # N504) in all test wells at 1:130,000 dilution for 1 hour at room temperature and then washed and incubated with 100 µL per well of TMB substrate (Product # 34028) for 30 minutes at room temperature in the dark. The plate was stopped with 0.16M sulfuric acid (Product # N600). Absorbances were read on a spectrophotometer at 450-550 nm.
- Conjugate
- Biotin
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of EGF (green) in HeLa cells either left untreated (left panel) or treated with 10 ng/mL EGF (right panel) for 5 minutes. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA (Product # 37525) for 15 minutes at room temperature. Cells were probed with an EGF monoclonal antibody (Product # M806) at a dilution of 1:50 for at least 1 hour at room temperature, washed with PBS, and incubated with DyLight 488 goat anti-mouse IgG secondary antibody (Product # 35502) at a dilution of 1:400 for 30 minutes at room temperature. F-Actin (red) was stained with DyLight 554 Phalloidin (Product # 21834) and nuclei (blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ArrayScan at 20X magnification.
- Conjugate
- Biotin