Invitrogen

PAX3 Polyclonal Antibody

Product Image Gallery

PAX3 Antibody — Fig. 4 Neuroprogenitors are generated normally in Ntn1 , Dcc , and Robo3 knockouts. a Immunohistochemistry of phospho-Histone H3, a mitotic marker, Ki67, a cell proliferation marker, SOX2, a neuroprogenitor marker, and PAX3/7, a dorsal progenitor marker in E10.5 spinal cord. b Quantification of phenotypes in (a). Data are normalized to WT and are represented as the mean +- SEM (Student and apos;s t -test; ns, not significant). Scale bar, 50 mum

PAX3 Antibody in Immunofluorescence (IF) — Immunofluorescence was performed on mouse E10.5 embryo trunk at the forelimb level. Embryos were fixed with 4%PFA in 0.1M phosphate buffer, pH 7.2 at 4C for 30 minutes, processed for cryoprotection in 20% sucrose, and embedded in OCT compound for cryosectioning at 10 µm thickness. Following sectioning, tissues were washed in PBS with 0.1% Triton-X 100 (PBT), blocked in 1% goat serum in PBT for 30 minutes at room temperature, and then probed with a Pax3 polyclonal antibody (Product # PA1-107) at a concentration of 1 µg/mL in blocking buffer overnight at 4C. Tissues were washed extensively with PBT, and detection was performed using an Alexa Fluor 568-conjugated goat anti-rabbit IgG secondary antibody (red) at a dilution of 1:1000 in blocking buffer. Nuclei (blue) were stained with DAPI. Note: Staining was observed in the dorsal neural tube (symmetric two halves on the left side of the image), the dermomyotome/myotome (middle streak between symmetric halves), and limb muscle progenitors (scattered cells to the right side of the image). Data supplied courtesy of the Innovator's Program.

PAX3 Antibody in Western Blot (WB) — Western blot analysis of Pax3 was performed by loading the indicated amounts of Pax3 and Pax4 (negative control) recombinant proteins, and 10 µL of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a Novex® 4-20% Tris-Glycine polyacrylamide gel. Proteins were transferred to a PVDF membrane using the G2 Fast Blotter (Product # 62288), and blocked with 5% milk in TBST for at least 1 hour at room temperature. Pax3 was detected at ~53 kD using a Pax3 polyclonal antibody (Product # PA1-107) at a dilution of 1:1000 in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-rabbit IgG secondary antibody (Product # 31460) at a dilution of 1:40,000 for at least 30 minutes. Chemiluminescent detection was performed using SuperSignal West Pico (Product # 34080). NOTE: The presence of Pax4 recombinant protein was determined by probing a duplicate blot with a Pax4 polyclonal antibody (Product # PA1-108), and the corresponding image is on the webpage for Product # PA1-108.

PAX3 Antibody in Immunoprecipitation (IP) — Immunoprecipitation of Pax3 was performed on RIPA cell extracts originating from a 6cm dish of HeLa cells transfected for 24 hours with a Pax3-HA construct. Cells were lysed in 1 mL of RIPA buffer, and antigen-antibody complexes were formed by incubating 225 µL of HeLa Pax-HA cell extract with 2 µg of a Pax3 polyclonal antibody (Product # PA1-107) and Protein A sepharose for 4 hours at 4C. The samples were washed extensively with RIPA buffer, and eluted by boiling in SDS sample buffer. Samples were resolved on an 8% Tris-Glycine gel alongside whole cell lysate (WCL) from empty vector and Pax3-HA-transfected HeLa cells, transferred to a nitrocellulose membrane, and blocked with 2% BSA in PBST for 10 minutes at room temperature. The membrane was probed with an anti-HA polyclonal antibody overnight at 4C, washed in TBST, and probed with an HRP-conjugated anti-rabbit IgG secondary antibody. Chemiluminescent detection was performed using ECL substrate. Data supplied courtesy of the Innovator's Program.

Product Details

Tested Applications

Dilution

Immunocytochemistry (ICC)

1:100-1:1000

Immunofluorescence (IF)

1:100-1:1000

Immunohistochemistry (Frozen) (IHC (F))

1:100-1:1000

Immunoprecipitation (IP)

2 µg

Western Blot (WB)

1:500-1:1000

Published Applications

Immunohistochemistry (IHC)

See 2 publications below

Immunofluorescence (IF)

See 2 publications below

Product Specifications

Species Reactivity

Human, Mouse

Published species

Mouse

Host / Isotype

Rabbit / IgG

Class

Polyclonal

Type

Antibody

Immunogen

Amino acids 301-442 of human PAX3

Conjugate

Unconjugated

Form

Liquid

Concentration

1 mg/mL

Purification

Protein G

Storage buffer

PBS with 1mg/mL BSA, 30% glycerol

Contains

0.05% sodium azide

Storage conditions

-20°C

RRID

AB_2539881

Product Specific Information

PA1-107 has successfully been used in Western blot for detecting overexpressed and recombinant human Pax3 protein, but may not be suitable for detecting endogenous Pax3.

By immunofluorescence, PA1-107 has been used to stain frozen embryonic mouse brain tissue.

PA1-107 has been validated for IP using overexpressed Pax3 protein.

Target Information

Pax 3 (Paired Box 3) is a transcription factor that is involved in development of the peripheral nervous system, melanocytes, some vascular smooth muscle, and a number of other derivatives. It regulates neurogenesis in pre-migratory neural crest cells from the dorsal neural tube, and in myogenic progenitors in the presomitic mesoderm and the hypaxial somites. Alternative splicing results in isoforms having different C-termini.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Bioinformatics

Protein Aliases: CDHS; HUP2; Paired Box 3; Paired Box Gene 3; Paired Box Homeotic Gene 3; Paired box protein Pax-3; Paired Box Protein-3; Paired Domain Gene 3; Paired Domain Gene HuP2; PAX3/FKHR fusion; Waardenburg Syndrome 1; WS1; WS3

Gene Aliases: CDHS; HUP2; Pax-3; PAX3; Sp; splotch; WS1; WS3

UniProt ID: (Human) P23760, (Mouse) P24610

Entrez Gene ID: (Human) 5077, (Mouse) 18505

Molecular Function: DNA binding protein helix-turn-helix transcription factor homeobox transcription factor nucleic acid binding transcription factor