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Western blotAntibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
- Immunocytochemistry [1]
- Chromatin Immunoprecipitation [1]
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- Product number
- 710178 - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- PAX3 Recombinant Polyclonal Antibody (16HCLC)
- Antibody type
- Polyclonal
- Antigen
- Recombinant full-length protein
- Description
- This antibody is predicted to react with mouse based on sequence homology.
- Antibody clone number
- 16HCLC
- Concentration
- 0.5 mg/mL
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis was performed on whole cell extracts of HeLa (Lane 1) and A431 (Lane 2). The blot was probed with Anti-PAX3 Recombinant Rabbit Polyclonal Antibody (Product # 710178, 2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 53 kDa band corresponding to PAX3 was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex®NuPAGE®12 % Bis-Tris gel (Product # NP0342BOX), XCell SureLock Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody. Chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Western blot analysis of PAX3 in A431 whole cell extracts using a PAX3 Recombinant Rabbit Polyclonal Antibody (Product # 710178) at a dilution of 1 µg/mL. Samples were detected using chemiluminescence (ECL). Results show a band at ~53kDa.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Immunofluorescent analysis of PAX3 in HeLa cells using a PAX3 Recombinant Rabbit Polyclonal Antibody (Product # 710178) followed by detection using an Alexa Fluor 488-conjugated Goat anti-Rabbit secondary antibody (green) (Image A). Nuclei were stained using DAPI (Image B) and actin stained with Alexa Fluor 594 phalloidin (red) (image C). Image D is a composite image showing nuclear localization of Pax 3.
Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Chromatin immunoprecipitation analysis of Pax3 was performed using cross-linked chromatin from 1 x 10^6 HCT116 human colon carcinoma cells treated with serum for 0, 15, and 60 minutes. Immunoprecipitation was performed using a multiplex microplate Matrix ChIP assay (see reference for Matrix ChIP protocol: http://www.ncbi.nlm.nih.gov/pubmed/22098709) with 1.0 µL/100 µL well volume of a Pax3 Recombinant Rabbit Polyclonal Antibody (Product # 710178). Chromatin aliquots from ~1 x 10^5 cells were used per ChIP pull-down. Quantitative PCR data were done in quadruplicate using 1 µL of eluted DNA in 2 µL SYBR real-time PCR reactions containing primers to amplify-3.2kb upstream of human FOS, exon-4 of FOS, or the imprinting control region (ICR) of the human H19 locus. PCR calibration curves were generated for each primer pair from a dilution series of sheared total genomic DNA. Quantitation of immunoprecipitated chromatin is presented as signal relative to the total amount of input chromatin. Results represent the mean +/- SEM for three experiments. A schematic representation of human FOS and H19 loci are shown above the data where boxes represent exons (grey boxes = translated regions, white boxes = untranslated regions), the zigzag lines represent introns, and the straight lines represent upstream sequences. Regions amplified by FOS and H19 primers are represented by black bars. Data courtesy of the Innovators Program.
