Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [3]
- Immunocytochemistry [2]
- Immunohistochemistry [2]
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Validation data
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- Product number
- PA5-27408 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- NONO Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Recombinant protein fragment
- Description
- Recommended positive controls: 293T, A431, H1299, HeLa, HepG2, Molt-4, Raji. Predicted reactivity: Mouse (100%), Rat (100%), Xenopus laevis (87%), Chicken (90%), Chimpanzee (100%), Bovine (99%). Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.
- Reactivity
- Human, Mouse
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µL
- Concentration
- 0.72 mg/mL
- 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 using NONO Polyclonal Antibody (Product # PA5-27408). Sample (30 µg of whole cell lysate). Lane A: Hep G2. 10% SDS PAGE. NONO Polyclonal Antibody (Product # PA5-27408) diluted at 1:1,000.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Knockdown of Non-POU domain-containing octamer-binding protein was achieved by transfecting HeLa with Non-POU domain-containing octamer-binding protein specific siRNAs (Silencer® select Product # s9613, s9614). Western blot analysis (Fig. a) was performed using Nuclear enriched extracts from the Non-POU domain-containing octamer-binding protein knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with NONO Polyclonal Antibody (Product # PA5-27408, 1:1000 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:6000 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 Non-POU domain-containing octamer-binding protein.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-NONO Polyclonal Antibody (Product # PA5-27408) and a 54kDa band corresponding to Non-POU domain-containing octamer-binding protein was observed across cell lines tested. Nuclear enriched extracts (30 µg lysate) of HeLa (Lane 1), K-562 (Lane 2), Hep G2 (Lane 3), PANC-1 (Lane 4), MCF7 (Lane 5), NTERA-2 cl.D1 (Lane 6), HEK-293 (Lane 7), NIH/3T3 (Lane 8) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0321BOX). 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 (1:2500 dilution) and detected by chemiluminescence with Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:6000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunocytochemistry-Immunofluorescence analysis of NONO was performed in HeLa cells fixed in 4% paraformaldehyde at RT for 15 min. Green: NONO Polyclonal Antibody (Product # PA5-27408) diluted at 1:500. Red: alpha Tubulin, a cytoskeleton marker. Blue: Hoechst 33342 staining.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescence analysis of Non-POU domain-containing octamer-binding protein 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 15 minutes, and blocked with 2% BSA for 45 minutes at room temperature. The cells were labeled with NONO Polyclonal Antibody (Product # PA5-27408) at 1:500 dilution in 0.1% BSA, incubated at 4 degree celsius overnight and then labeled with Donkey anti-Rabbit IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, Alexa Fluor Plus 488 (Product # A32790), (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). Panel d represents the merged image showing nuclear 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)
- Main image
- Experimental details
- Immunohistochemistry (Paraffin) analysis of NONO was performed in paraffin-embedded mouse brain tissue using NONO Polyclonal Antibody (Product # PA5-27408) at a dilution of 1:500.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunohistochemical analysis of paraffin-embedded Cal27 xenograft, using p54/nrb (Product # PA5-27408) antibody at 1:100 dilution. Antigen Retrieval: EDTA based buffer, pH 8.0, 15 min.