Invitrogen

PAF49 Polyclonal Antibody

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PAF49 Antibody in Immunocytochemistry (ICC/IF) — Immunocytochemistry-Immunofluorescence analysis of PAF49 was performed in HeLa cells fixed in ice-cold MeOH for 5 min. Green: PAF49 Polyclonal Antibody (Product # PA5-27632) diluted at 1:1000. Red: alpha Tubulin, a cytoskeleton marker. Blue: Hoechst 33342 staining.

PAF49 Antibody in Immunohistochemistry (Paraffin) (IHC (P)) — Immunohistochemistry (Paraffin) analysis of PAF49 was performed in paraffin-embedded human ovarian cancer tissue using PAF49 Polyclonal Antibody (Product # PA5-27632) at a dilution of 1:500.

PAF49 Antibody in Western Blot (WB) — Western Blot analysis of PAF49 was performed by separating 30 µg of various whole cell extracts by 10% SDS-PAGE. Proteins were transferred to a membrane and probed with a PAF49 Polyclonal Antibody (Product # PA5-27632) at a dilution of 1:1000.

PAF49 Antibody in Immunoprecipitation (IP) — Immunoprecipitation of CD3EAP was performed in K562 cells. Antigen-antibody complexes were formed by incubating approximately 500 µg whole cell lysate with 5 to 10 µL of polyclonal CD3EAP antibody (Product # PA5-27632) rotating 60 min at RT. The immune complexes were captured on 625 µg of anti-rabbit coated Dynabeads (Product # 11204D) and washed extensively. They were then eluted and analyzed using the Simple Western system using the same antibody as used in immunoprecipitation at a dilution of 1:25, followed by a 1:100 dilution of secondary antibody. Lane 1 is the input, lane 2 no antibody IP and lane 3 is the target specific IP. Data courtesy of the Yeo lab as part of the ENCODE project.

PAF49 Antibody in ChIP assay (ChIP) — Chromatin immunoprecipitation analysis of PAF49 was performed using cross-linked chromatin from 1x10^6 HCT116 colon carcinoma cells treated with serum for 0 and 30 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 PAF49 polyclonal antibody (Product # PA5-27632). Chromatin aliquots from ~1x10^5 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 exon-1 of the RNA5-8S5 gene or -20 kb upstream of the LAMC1 gene. 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 representations of the RNA5-8S5 and LAMC1 loci are shown above the data where boxes represent exons (black boxes = translated regions, white boxes = untranslated regions), the zigzag line represents an intron, and the straight line represents upstream sequence. Regions amplified by RNA5-8S5 and LAMC1 primers are represented by black bars. Data courtesy of the Innovators Program.

PAF49 Antibody in RNA Immunoprecipitation (RIP) — RNA immunoprecipitation (RIP) western of CD3EAP was performed in K562 cells. Antigen-antibody complexes were formed by incubating approximately 500 µg whole cell lysate with 5 to 10 µL of polyclonal CD3EAP antibody (Product # PA5-27632) rotating 60 min at RT. The immune complexes were captured on 625 µg of anti-rabbit coated Dynabeads (Product # 11204D) and washed extensively. They were then eluted and analyzed using the Simple Western system using the same antibody as used in immunoprecipitation at a dilution of 1:25, followed by a 1:100 dilution of secondary antibody. Lane 1 is the input, lane 2 no antibody IP and lane 3 is the target specific IP. Data courtesy of the Yeo lab as part of the ENCODE project.

Product Details

PA5-27632

Applications	Tested Dilution	Publications
Western Blot (WB)	1:500-1:3,000	-
Immunohistochemistry (Paraffin) (IHC (P))	1:100-1:1,000	-
Immunocytochemistry (ICC/IF)	1:100-1:1,000	-
ChIP assay (ChIP)	Assay-Dependent	-
RNA Immunoprecipitation (RIP)	Assay-dependent	-

Product Specifications
Species Reactivity	Human
Host/Isotype	Rabbit / IgG
Class	Polyclonal
Type	Antibody
Immunogen	Recombinant fragment corresponding to a region within amino acids 1 and 268 of Human PAF49 View antigen
Conjugate	Unconjugated
Form	Liquid
Concentration	0.55 mg/mL
Purification	Antigen affinity chromatography
Storage buffer	PBS, pH 7, with 20% glycerol
Contains	0.025% ProClin 300
Storage conditions	Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.
RRID	AB_2545108

Product Specific Information

Recommended positive controls: A549, H1299, HCT116.

Store product as a concentrated solution. Centrifuge briefly prior to opening the vial.

Target Information

DNA-dependent RNA polymerase catalyzes the transcription of DNA into RNA using the four ribonucleoside triphosphates as substrates. Component of RNA polymerase I which synthesizes ribosomal RNA precursors. Isoform 1 is involved in UBTF-activated transcription, presumably at a step following PIC formation.; Isoform 2 has been described as a component of preformed T-cell receptor (TCR) complex. [UniProt]

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

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Bioinformatics

Protein Aliases: A34.5; antis; Antisense to ERCC-1 protein; ASE-1; CD3-epsilon-associated protein; CD3e antigen, epsilon polypeptide associated protein; CD3e molecule, epsilon associated protein; CD3E-associated protein; DNA-directed RNA polymerase I subunit G; DNA-directed RNA polymerase I subunit RPA34; RNA polymerase I-associated factor PAF49

Gene Aliases: ASE-1; ASE1; CAST; CD3EAP; PAF49; POLR1G

UniProt ID: (Human) O15446

Entrez Gene ID: (Human) 10849

Molecular Function: DNA-directed RNA polymerase