WB
Western blot analysis of Dnmt3b on NCCIT cell lysates with Rabbit anti-Dnmt3b antibody at 1/2,000 dilution. Lysates/proteins at 20 µg/Lane. Exposure time: 1 minute 2 seconds; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.ICC/IF
Immunocytochemistry analysis of NCCIT cells labeling Dnmt3b with Rabbit anti-Dnmt3b antibody at 1/250 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Dnmt3b antibody at 1/250 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (594) was used as the secondary antibody at 1/1,000 dilution.ChIP
Chromatin immunoprecipitations were performed with cross-linked chromatin from NCCIT cells with Dnmt3b or Normal Rabbit IgG according to the ChIP protocol. The enriched DNA was quantified by real-time PCR using indicated primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.| Product Name | Dnmt3b Recombinant Rabbit Monoclonal Antibody |
|---|---|
| Antibody Type | Primary Antibodies |
| Product description | Methylation at the 5'-position of cytosine is the only known naturally occurring covalent modification of the mammalian genome. DNA methylation requires the enzymatic activity of DNA 5-cytosine methyltransferase (Dnmt) proteins, which catalyze the transfer of a methyl group from S-adenosyl methionine to the 5'-position of cytosines residing in the dinucleotide CpG motif, and this methylation results in transcriptional repression of the target gene. The Dnmt enzymes are encoded by independent genes. Dnmt1 is the most abundant, and it preferentially methylates hemimethylated DNA and coordinates gene expression during development. Additional mammalian Dnmt proteins include Dnmt2 and Dnmt3. Dnmt2 lacks the large N-terminal regulator domain of Dnmt1, is expressed at substantially lower levels in adult tissues, and is likely involved in methylating newly integrated retroviral DNA. Dnmt3a and Dnmt3b are encoded by two distinct genes, but both are abundantly expressed in embryonic stem cells, where they also methylate CpG motifs on DNA. |
| Immunogen | Synthetic peptide within Human Dnmt3b aa 61-110 / 853. |
| Clonality | Monoclonal |
|---|---|
| Isotype | IgG |
| Host Species | Recombinant rabbit |
| Tested Applications | ChIPICC/IFWB |
| WB:1:2000 ICC/IF:1:50-1:250 ChIP:Use 2 μg for 25 μg of chromatin. |
|
| Species Reactivity | HumanRat |
| Concentration | 1mg/ml |
| Purification | Protein A |
| Alternative Names | Cytosine 5methyltransferase 3B antibody DNA antibody DNA (cytosine 5) methyltransferase 3 beta antibody DNA (cytosine 5)-methyltransferase 3B antibody DNA (cytosine-5)-methyltransferase 3B antibody DNA methyltransferase HsaIIIB antibody DNA MTase HsaIIIB antibody DNM3B_HUMAN antibody Dnmt3b antibody EC 2.1.1.37 antibody ICF antibody ICF1 antibody M.HsaIIIB antibody MGC124407 antibody RP23-89H14.3 antibody |
|---|---|
| Molecular Weight(MW) | 96 kDa(Observed band size: 81 kDa) |
| Cellular Localization | Nucleus. |
WB
Western blot analysis of Dnmt3b on NCCIT cell lysates with Rabbit anti-Dnmt3b antibody at 1/2,000 dilution. Lysates/proteins at 20 µg/Lane. Exposure time: 1 minute 2 seconds; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.
ICC/IF
Immunocytochemistry analysis of NCCIT cells labeling Dnmt3b with Rabbit anti-Dnmt3b antibody at 1/250 dilution. Cells were fixed in 4% paraformaldehyde for 15 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 15 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Dnmt3b antibody at 1/250 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (488) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI. Beta tubulin (red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (594) was used as the secondary antibody at 1/1,000 dilution.
ChIP
Chromatin immunoprecipitations were performed with cross-linked chromatin from NCCIT cells with Dnmt3b or Normal Rabbit IgG according to the ChIP protocol. The enriched DNA was quantified by real-time PCR using indicated primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.| Application Notes | WB:1:2000 ICC/IF:1:50-1:250 ChIP:Use 2 μg for 25 μg of chromatin. |
|---|
| Form | Liquid |
|---|---|
| Storage Instructions | Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles. |
| Storage Buffer | PBS (pH7.4), 0.1% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide. |
