Antibody data
- Antibody Data
- Antigen structure
- References [1]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [5]
- Immunohistochemistry [2]
- Other assay [2]
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Validation data
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- Product number
- MA1-10050 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- AHNAK Monoclonal Antibody (EM-09)
- Antibody type
- Monoclonal
- Antigen
- Other
- Description
- This antibody reacts with AHNAK1, a 700 kDa multi-functional adaptor protein expressed mainly in epithelial cell, various types of muscle cells and immune cells. Immunocytochemistry: permeabilization is required.
- Reactivity
- Human, Mouse
- Host
- Mouse
- Isotype
- IgG
- Antibody clone number
- EM-09
- Vial size
- 100 µg
- Concentration
- 1 mg/mL
- Storage
- 4° C, do not freeze
Submitted references A yeast display immunoprecipitation screen for targeted discovery of antibodies against membrane protein complexes.
Lajoie JM, Cho YK, Frost D, Bremner S, Li L, Shusta EV
Protein engineering, design & selection : PEDS 2019 Dec 31;32(5):219-230
Protein engineering, design & selection : PEDS 2019 Dec 31;32(5):219-230
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of AHNAK was achieved by transfecting HaCaT with AHNAK specific siRNAs (Silencer® select Product # S41427, S41428). Western blot analysis (Fig. a) was performed using whole cell extracts prepared with RIPA buffer, from the AHNAK knockdown cells (lane 2) and non-targeting scrambled siRNA transfected cells (lane 1). The blot was probed with AHNAK Monoclonal Antibody (EM-09) (Product # MA1-10050, 1:2000) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:20,000) and detected by chemiluminescence using the iBright™ FL1500 Imaging System (Product # A44115). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to AHNAK.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using AHNAK Monoclonal Antibody (EM-09) (Product # MA1-10050) and a 680 kDa band corresponding to AHNAK was observed across cell lines tested. Whole cell extracts (60 µg lysate), prepared with RIPA buffer, of U-87 MG (Lane 1), PANC-1 (Lane 2), 769-P (Lane 3), NIH:OVCAR-3 (Lane 4), HaCaT (Lane 5), SW480 (Lane 6), Jurkat (Lane 7), A-673 (Lane 8), HEL 92.1.7 (Lane 9) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX), 10 well. Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23002) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:2000) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:20,000) using the iBright™ FL1500 Imaging System (Product # A44115). Chemiluminescent detection was performed using SuperSignal™ West Atto Ultimate Sensitivity Substrate (Product # A38556). As reported, the levels of AHNAK were low in Jurkat, A-673 and HEL.92.17 cell lines as shown here in the figure.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence staining of AHNAK1 in human primary fibroblasts using anti-AHNAK1 (EM-09; green). Actin filaments were decorated by phalloidin (red) and cell nuclei stained with DAPI (blue).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence staining of AHNAK1 in HeLa cell line using anti-AHNAK1 (EM-09; red). Cell nuclei stained with DAPI (blue).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemistry staining of AHNAK1 in HeLa cell line using anti-AHNAK1 (EM-09; red) Monoclonal antibody (Product # MA1-10050). Cell nuclei stained with DAPI (blue).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of AHNAK was performed using 70% confluent log phase HaCaT cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 15 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with AHNAK Monoclonal Antibody (EM-09) (Product # MA1-10050, 1:100) in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor™ Plus 488 (Product # A32766, 1:2000), for 45 minutes at room temperature (Panel a: Green). Nuclei (Panel b:Blue) were stained with ProLong™ Diamond Antifade Mountant with DAPI (Product # P36962). F-actin (Panel c: Red) was stained with Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cell membrane and cytosolic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemistry staining of AHNAK1 in primary human fibroblasts using anti-AHNAK1 (EM-09; green) Monoclonal antibody (Product # MA1-10050). Actin filaments were decorated by phalloidin (red) and cell nuclei stained with DAPI (blue).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry staining (frozen sections) of murine tongue by anti-AHNAK1 antibody (EM-09; red). Actin filaments were decorated by phalloidin (green), cell nuclei stained with DAPI (blue).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemistry staining (frozen sections) of murine tongue by anti-AHNAK1 (EM-09; red) Monoclonal antibody (Product # MA1-10050). Actin filaments were decorated by phalloidin (green), cell nuclei stained with DAPI (blue).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- NULL
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- NULL