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Western blot
ELISAAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [4]
- Immunocytochemistry [1]
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- Product number
- MA5-15494 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Cytokeratin 5 Monoclonal Antibody (1E1)
- Antibody type
- Monoclonal
- Antigen
- Purifed from natural sources
- Description
- MA5-15494 targets Cytokeratin 5 in indirect ELISA, WB applications and shows reactivity with Human samples. The MA5-15494 immunogen is purified recombinant fragment of Cytokeratin 5 expressed in E. Coli. . MA5-15494 detects Cytokeratin 5 which has a predicted molecular weight of approximately 62.3kDa.
- Reactivity
- Human, Mouse
- Host
- Mouse
- Isotype
- IgG
- Antibody clone number
- 1E1
- Vial size
- 100 µL
- Concentration
- Conc. Not Determined
- 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 of Cytokeratin 5 using Cytokeratin 5 monoclonal antibody (Product # MA5-15494) in HeLa cell lysate (1).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of Cytokeratin 5 using Cytokeratin 5 monoclonal antibody (Product # MA5-15494) in HeLa cell lysate (1).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of Cytokeratin 5 was achieved by transfecting A-431 with Cytokeratin 5 specific siRNAs (Silencer® select Product # s7956, S7957). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Cytokeratin 5 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with Cytokeratin 5 Monoclonal Antibody (1E1) (Product # MA5-15494, 1:1000 dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution). 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 Cytokeratin 5.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-Cytokeratin 5 Monoclonal Antibody (1E1) (Product # MA5-15494) and a 60kDa band corresponding to Cytokeratin 5 was observed across the cell lines tested except MCF7, IMR-32 and SH-SY5Y along with non specific bands around 40-55 kDa. Whole cell extracts (30 µg lysate) of MCF 10A (Lane 1), MCF7 (Lane 2), A-431 (Lane 3), HaCaT (Lane 4), IMR-32 (Lane 5) and SH-SY 5Y (Lane 6), were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # LC2002) by using iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000 dilution) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Cytokeratin 5 was performed using 70% confluent log phase A-431 cells. The cells were fixed with ice-cold acetone at 4°C for 15 minutes and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with Cytokeratin 5 Monoclonal Antibody (1E1) (Product # MA5-15494) at 1:100 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175), (1:2000 dilution), 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 dilution). Panel d represents the merged image showing cytoskeletal localization. Panel e represents MCF7 cells having lower expression levels of KRT5. Panel f represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.
