- MA5-27399 - Invitrogen Antibodies CCDC22 antibody

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of CCDC22 in HEK293T cells in untransfected (Left lane) and transfected (Right lane) samples using 5 µg per lane. The samples were separated by SDS-PAGE and probed with CCDC22 (Product # MA5-27399) monoclonal antibody.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Knockdown of Coiled-coil domain-containing protein 22 was achieved by transfecting NIH:OVCAR-3 with Coiled-coil domain-containing protein 22 specific siRNAs (Silencer® select Product # S226233, S26283). Western Blot analysis (Fig. a) was performed using Whole cell extracts from the Coiled-coil domain-containing protein 22 knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with CCDC22 Monoclonal Antibody (OTI5C1) (Product # MA5-27399, 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 Coiled-coil domain-containing protein 22.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western Blot was performed using Anti-CCDC22 Monoclonal Antibody (OTI5C1) (Product # MA5-27399) and a 70 kDa band corresponding to Coiled-coil domain-containing protein 22 was observed across all the tested cell lines. Whole cell extracts (40 µg lysate) of NIH:OVCAR-3 (Lane 1), HEK-293 (Lane 2), SK-O-V3 (Lane 3) were electrophoresed using NuPAGE™ 10% Bis-Tris Protein Gel (Product # NP0301BOX). 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 (1:1000 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)
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Experimental details: Immunofluorescence analysis of Coiled-coil domain-containing protein 22 was performed using 70% confluent log phase NIH:OVCAR-3 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 CCDC22 Monoclonal Antibody (OTI5C1) (Product # MA5-27399) at 5 µg/mL 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 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 cytoplasmic localization. Panel e represents control cells with no primary antibody to assess background. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry was performed on paraffin-embedded human endometrium tissue. To expose target proteins, heat-induced epitope retrieval by 10mM citric buffer, pH6.0, 100°C for 10min. Following antigen retrieval, tissues were probed with a CCDC22 monoclonal antibody (Product # MA5-27399) at a dilution of 1:500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemistry was performed on paraffin-embedded human prostate tissue. To expose target proteins, heat-induced epitope retrieval by 10mM citric buffer, pH6.0, 100°C for 10min. Following antigen retrieval, tissues were probed with a CCDC22 monoclonal antibody (Product # MA5-27399) at a dilution of 1:500.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry was performed on paraffin-embedded carcinoma of human prostate tissue. To expose target proteins, heat-induced epitope retrieval by 10mM citric buffer, pH6.0, 100°C for 10min. Following antigen retrieval, tissues were probed with a CCDC22 monoclonal antibody (Product # MA5-27399) at a dilution of 1:500.

MA5-27399