Antibody data
- Antibody Data
- Antigen structure
- References [0]
- Comments [0]
- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Immunohistochemistry [1]
- Chromatin Immunoprecipitation [2]
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Validation data
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- Product number
- PA5-32443 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- HDAC1 Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Description
- Heat-mediated antigen retrieval is recommended prior to staining, using a 10mM citrate buffer, pH 6.0, for 10 minutes followed by cooling at room temperature for 20 min. Following antigen retrieval, incubate samples with primary antibody for 10 min at room temperature. A suggested positive control is tonsil or breast carcinoma.
- Reactivity
- Human, Mouse
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 500 µL
- 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
- Knockdown of HDAC1 was achieved by transfecting MCF7 with HDAC1 specific siRNAs (Silencer® select Product # s73). Western blot analysis (Fig. a) was performed using nuclear enriched cell extracts (20ug) from the HDAC1 knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with HDAC1 Polyclonal Antibody (Product # PA5-32443, 1:2000 dilution) 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 HDAC1.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot was performed using Anti-HDAC1 Polyclonal Antibody (Product # PA5-32443) and a 58kDa band corresponding to HDAC1 was observed in all cell lines tested. Modified whole cell extracts (1% SDS) (30 µg lysate) of MCF7 (Lane 1), HeLa (Lane 2), HEK-293 (Lane 3) and NIH/3T3 (Lane 4) were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) 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-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
- Immunofluorescence analysis of HDAC 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 1 hour at room temperature. The cells were labeled with HDAC1 Polyclonal Antibody (Product # PA5-32443) at 1:100 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) at a dilution of 1:2000 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). 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
- Immunohistochemical analysis of HDAC1 using anti-HDAC1 Polyclonal Antibody (Product # PA5-32443) in Breast Carcinoma Cancer Tissue. The recommened dilution for this antibody in immunohistochemistry applications is 1:200.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Chromatin Immunoprecipitation (ChIP) assay of endogenous HDAC1 protein using HDAC1 Polyclonal Antibody: ChIP was performed using HDAC1 Polyclonal Antibody (Product # PA5-32443, 5 µg) on sheared chromatin from MCF-7 cells using the MAGnify ChIP System kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to CDKN1-PR, GAPDH- TSS and FOSL1-PR as active binding regions and FOSL1- Exon1 and SAT alpha as inactive binding regions. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method. PR: Promoter, TSS: Transcription start site.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Chromatin Immunoprecipitation (ChIP) assay of endogenous HDAC1 protein using Anti-HDAC1 Antibody: ChIP was performed using Anti-HDAC1 Rabbit Polyclonal Antibody (Product # PA5-32443, 5 µg) on sheared chromatin from HeLa cells using the MAGnify ChIP System kit (Product # 49-2024). Normal Rabbit IgG was used as a negative IP control. The purified DNA was analyzed by qPCR using primers binding to CDKN1A Intron 1, transcriptional start site and gene body (+2kb) of GAPDH, RPL30 exon 5 and SATA satellite repeats. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.