PA5-19975

antibody from Invitrogen Antibodies

Targeting: DIABLO DFNA64, DIABLO-S, FLJ10537, FLJ25049, SMAC

Immunoprecipitation

Antibody data

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Validation data

Product number: PA5-19975 - Provider product page
Provider: Invitrogen Antibodies
Product name: DIABLO Polyclonal Antibody
Antibody type: Polyclonal
Antigen: Synthetic peptide
Description: In Western blot applications, this antibody detects a band at ~25kDa. A suggested positive control is mouse heart tissue lysate. PA5-19975 can be used with blocking peptide PEP-0100.
Reactivity: Human, Mouse, Rat
Host: Rabbit
Isotype: IgG
Vial size: 100 µg
Concentration: 1 mg/mL
Storage: Maintain refrigerated at 2-8°C for up to 3 months. For long term storage store at -20°C

DIABLO protein structure - PA5-19975 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot Validation in (A and B) Mouse Heart Tissue Lysate and (C) Rat Heart Tissue Lysate. Loading: 15 µg of lysates per lane. Antibodies: DIABLO Polyclonal Antibody (Product # PA5-19975) (1 µg/mL), 1h incubation at RT in 0.05 NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10,000 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot Validation in (A and B) Mouse Heart Tissue Lysate and (C) Rat Heart Tissue Lysate. Loading: 15 µg of lysates per lane. Antibodies: DIABLO Polyclonal Antibody (Product # PA5-19975) (1 µg/mL), 1h incubation at RT in 0.05 NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10,000 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western Blot Validation in Human, Mouse and Rat Cell Lines. Loading: 15 µg of lysates per lane. Antibodies: DIABLO Polyclonal Antibody (Product # PA5-19975) (1 µg/mL), 1h incubation at RT in 0.05 NFDM/TBST. Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10,000 dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of DIABLO was achieved by transfecting HeLa cells with DIABLO specific siRNAs (Silencer® select Product # S32187, S32188). Western blot analysis (Fig. a) was performed using whole cell extracts from the DIABLO knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with DIABLO Polyclonal Antibody (Product # PA5-19975, 1 ug/ml) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 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 DIABLO.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot was performed using Anti-DIABLO Polyclonal Antibody(Product # PA5-19975) and a 21kDa band corresponding to DIABLO was observed across the cell lines and tissues tested. Whole cell extracts (30 µg lysate) of HeLa (Lane 1), MCF-7 (Lane 2), Hep G2 (Lane 3), Mouse Testis (Lane 4), Mouse Skeletal Muscle (Lane 5) Mouse Brain (Lane 6) and Mouse Heart (Lane 7) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # LC2001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1 ug/ml) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036,1:4000 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: Immunofluorescent analysis of mouse spleen cells using a Smac polyclonal antibody (Product # PA5-19975) at a 10 µg/mL dilution.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of 4% paraformaldehyde-fixed Mouse Spleen Cells labeling Smac with DIABLO Polyclonal Antibody (Product # PA5-19975) at 10 µg/mL, followed by goat anti-rabbit IgG secondary antibody at 1:500 dilution (red).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of DIABLO was performed using 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with DIABLO Polyclonal Antibody (Product # PA5-19975) at 5 µg/mL in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 1:2000 dilution), 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 dilution). Panel d represents the merged image showing Mitochondrial localization. Panel e represents control HeLa cells with no primary antibody to assess background. The images were captured at 60X magnification.