|Product Name||Anti-DUT Antibody|
|Storage & Handling||At -20°C for one year. After reconstitution, at 4°C for one month. It can also be aliquotted and stored frozen at -20°C for a longer time.Avoid repeated freezing and thawing.|
|Description||Rabbit IgG polyclonal antibody for Deoxyuridine 5'-triphosphate nucleotidohydrolase, mitochondrial(DUT) detection. Tested with WB, IHC-P in Human;Mouse;Rat.|
|Cite This Product||Anti-DUT Antibody (Boster Biological Technology, Pleasanton CA, USA, Catalog # PA1030-1)|
|Contents/Buffer||Each vial contains 5mg BSA, 0.9mg NaCl, 0.2mg Na2HPO4, 0.05mg Thimerosal, 0.05mg NaN3.|
|Immunogen||A synthetic peptide corresponding to a sequence at the C-terminus of human DUT(212-229aa KKGDRIAQLICERIFYPE), different from the related rat and mouse sequences by two amino acids.|
|Reactivity||Human, Mouse, Rat|
Assay Dilutions Overview
Immunohistochemistry(Paraffin-embedded Section), 0.5-1μg/ml, Human, Rat, Mouse, By Heat
Western blot, 0.1-0.5μg/ml, Human, Mouse, Rat
Boster's Secondary Antibodies And IHC, WB Kits
The following reagents are used to generate the images below.Boster recommends Enhanced Chemiluminescent Kit with anti-Rabbit IgG (EK1002) for Western blot, and HRP Conjugated anti-Rabbit IgG Super Vision Assay Kit (SV0002-1) for IHC(P).
Images And Assay Conditions
Figure 1. IHC analysis of DUT using anti-DUT antibody (PA1030-1).
DUT was detected in paraffin-embedded section of human intestinal cancer tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-DUT Antibody (PA1030-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.
Figure 2. Western blot analysis of DUT using anti-DUT antibody (PA1030-1).
Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions.
Lane 1: HELA whole cell lysates,
Lane 2: SMMC-7721 whole cell lysates.
After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-DUT antigen affinity purified polyclonal antibody (Catalog # PA1030-1) at 0.5 μg/mL overnight at 4°C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # EK1002) with Tanon 5200 system. A specific band was detected for DUT at approximately 27KD. The expected band size for DUT is at 27KD.
Figure 3. IHC analysis of DUT using anti-DUT antibody (PA1030-1).
DUT was detected in paraffin-embedded section of rat heart tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1μg/ml rabbit anti-DUT Antibody (PA1030-1) overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC)(Catalog # SA1022) with DAB as the chromogen.
Protein Target Info (Source: Uniprot.org)
|Protein Name||Deoxyuridine 5'-triphosphate nucleotidohydrolase, mitochondrial|
|Tissue Specificity||Found in a variety of tissues. Isoform 3 expression is constitutive, while isoform 2 expression correlates with the onset of DNA replication (at protein level). Isoform 2 degradation coincides with the cessation of nuclear DNA replication (at protein level). .|
|Alternative Names||Deoxyuridine 5'-triphosphate nucleotidohydrolase, mitochondrial;dUTPase;220.127.116.11;dUTP pyrophosphatase;DUT;|
|Subcellular Localization||Isoform 2: Nucleus.|
|Molecular Weight||26563 MW|
*if product is indicated to react with multiple species, protein info is based on the human gene.
|Protein Function||This enzyme is involved in nucleotide metabolism: it produces dUMP, the immediate precursor of thymidine nucleotides and it decreases the intracellular concentration of dUTP so that uracil cannot be incorporated into DNA. .|
|Research Areas||Dna / Nucleotides, Dna / Rna, Dna Synthesis, Epigenetics And Nuclear Signaling
*You can search these to find other products in these research areas.
|Background||Deoxyuridine triphosphate nucleotidohydrolase(dUTPase) is responsible for maintaining low intracellular levels of dUTP, thus preventing the incorporation of dUTP into DNA. dUTPase activity/expression can be down-regulated using siRNA specifically targeted to dUTPase mRNA and dUTPase plays a role in DNA nucleotide metabolism. This protein, present predominantly in the cytoplasm, contains 252 amino acids with a Mr of 26,704. It exhibits 35% identity with the E.coli dUTPase and 53% identity with the Saccharomyces cerevisiae enzyme. The nuclear and mitochondrial forms of dUTPase are encoded by the same gene with isoform-specific transcripts arising through the use of alternative 5-prime exons. Human dUTPase exhibits 92% identity with rat. Moreover, this enzyme has profound effects on the efficacy of agents that target thymidylate biosynthesis.|
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1. Post-translational modification:phosphorylation, methylation, glycosylation etc. These modifications prevent SDS molecules from binding to the target protein and thus make the band size appear larger than expected
2. Post-translational cleavage: this can cause smaller bands and or multiple bands
3. Alternative splicing: the same gene can have alternative splicing patterns generating different size proteins, all with reactivities to the antibody.
4. Amino Acid R chain charge: SDS binds to positive charges. The different size and charge of the Amino Acid side chains can affect the amount of SDS binding and thus affect the observed band size.
5. Multimers: Multimers are usually broken up in reducing conditions. However if the interactions between the multimers are strong, the band may appear higher.,