MGMT Recombinant Rabbit Monoclonal Antibody [JJ089-6] - BSA and Azide free
Catalog# HA752010
MGMT Recombinant Rabbit Monoclonal Antibody [JJ089-6] - BSA and Azide free
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WB
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IHC-P
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IF-Cell
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FC(Intra)
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Human
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ET1701-55
含抗保成分
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unconjugated
Overview
Product Name
MGMT Recombinant Rabbit Monoclonal Antibody [JJ089-6] - BSA and Azide free
Antibody Type
Recombinant Rabbit monoclonal Antibody
Immunogen
Recombinant protein within Human MGMT aa 1-140 / 207.
Species Reactivity
Human
Validated Applications
WB, IHC-P, IF-Cell, FC(Intra)
Target Molecular Weight
Predicted band size: 22 kDa
Positive Control
MCF7 cell lysate, Jurkat cell lysate, HCT 116 cell lysate, HeLa cell lysate, MCF7, human tonsil tissue.
Conjugation
unconjugated
Clone Number
JJ089-6
Product Features
Form
Liquid
Concentration
1 mg/mL.(The concentration of this product may be batch-dependent)
Storage Instructions
Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.
Storage Buffer
PBS (pH7.4).
Isotype
IgG
Purification Method
Protein A affinity purified.
Application Dilution
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WB
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1:2,000-1:5,000
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IHC-P
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1:50-1:200
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IF-Cell
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1:100
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FC(Intra)
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1:1,000
Target
Function
MGMT (O6-methylguanine-DNA methyltransferase) is transcriptionally activated in response to DNA damage and functions to repair mutagenic and cytotoxic O6-alkylguanine lesions caused by carcinogens or cytostatic drugs. MGMT induction by ionising radiation does not occur in p53-deficient mice, suggesting that MGMT induction may require p53. Similarly, MGMT mRNA and protein were shown to be inducible by ionising radiation, only in cell lines that express functional p53, and not in cell lines that do not express wild type p53. In contrast, high MGMT activity was associated with the presence of mutant p53, in a study of oral cancer cell lines. Similarly, MGMT activity was significantly lower in ovarian tumors with wildtype p53 than in tumors with mutant p53, supporting the view that wildtype p53 down-regulates the basal MGMT promoter.
Background References
1. Viel T et al. Optimizing Glioblastoma Temozolomide Chemotherapy Employing Lentiviral-based Anti-MGMT shRNA Technology. Mol Ther 21:570-9 (2013).
2. Viel T et al. Early assessment of the efficacy of temozolomide chemotherapy in experimental glioblastoma using [18F]FLT-PET imaging. PLoS One 8:e67911 (2013).
Sequence Similarity
Belongs to the MGMT family.
Subcellular Location
Nucleus.
UNIPROT
Synonyms
6 O methylguanine DNA methyltransferase antibody
6-O-methylguanine-DNA methyltransferase antibody
Agat antibody
AGT antibody
AI267024 antibody
EC 2.1.1.63 antibody
Methylated DNA protein cysteine methyltransferase antibody
Methylated-DNA--protein-cysteine methyltransferase antibody
Methylguanine DNA methyltransferase antibody
MGC107020 antibody
ExpandImages
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☑ Relative expression (RE)
Western blot analysis of MGMT on different lysates with Rabbit anti-MGMT antibody (HA752010) at 1/5,000 dilution.
Lane 1: MCF7 (Human breast cancer cell) cell lysate
Lane 2: MDA-MB-231 (Human breast cancer cell) cell lysate
Lane 3: Jurkat (Human T-lymphoblastic cells) cell lysate
Lane 4: HCT 116 (Human colon cancer cell) cell lysate
Lane 5: MOLT-4 (Human acute T-Cell leukemia cell) cell lysate
Lane 6: HeLa (Human cervical adenocarcinoma cell) cell lysate
Lane 7: HT-29 (Human colorectal adenocarcinoma cell) cell lysate
Lysates/proteins at 15 µg/Lane.
Exposure time: 25 seconds; ECL: K1801
MDA-MB-231 exhibits low MGMT expression (PMID: 26420420).
Blocking: 5% NFDM/TBST, 1 hour at room temperature
Primary antibody: HA752010, 1/5,000 in primary antibody dilution buffer (K1803), overnight at 4 ℃
Secondary antibody: Goat anti-Rabbit IgG-HRP (HA1001), 1/50,000 in 5% NFDM/TBST, 1 hour at room temperature
Predicted band size: 22 kDa
Observed band size: 22 kDa -
☑ Knockdown (KD)
Western blot analysis of MGMT on different lysates with Rabbit anti-MGMT antibody (HA752010) at 1/2,000 dilution.
Lane 1: HAP1-parental cell lysate
Lane 2: HAP1-MGMT KD cell lysate
Lysates/proteins at 10 µg/Lane.
Predicted band size: 22 kDa
Observed band size: 22 kDa
Exposure time: 2 minutes 40 seconds; ECL: K1801;
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 (HA752010) at 1/2,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Immunocytochemistry analysis of MCF7 cells labeling MGMT with Rabbit anti-MGMT antibody (HA752010) at 1/100 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-MGMT antibody (HA752010) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) 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 (HA601187, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
Application: Flow Cytometry (Intra)
Species: Human
Sample: MCF7 (Human breast cancer cell)
Fixation: 4% Paraformaldehyde, 15 minutes at room temperature.
Permeabilization: 0.1% Tween-20, 15 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 15 minutes at room temperature.
Antibody dilution buffer: 1x PBS.
Primary antibody: HA752010(1 μg/mL, Red) compared with Rabbit IgG Isotype Control (HA722127, Green), 15 minutes at room temperature.
Secondary antibody: Goat Anti-Rabbit IgG (iFluor™ 488, HA1121), 15 minutes at room temperature. -
Application: Immunohistochemistry (IHC-P)
Species: Human
Tissue: Tonsil
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA752010, 1/200, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature. -
Application: Immunohistochemistry (IHC-P)
Species: Human
Tissue: Lung carcinoma
Sample: Paraffin-embedded section
Antigen retrieval: Heat-mediated, Tris-EDTA buffer (pH 9.0), 20 minutes at 95℃.
Wash buffer: 1× TBST
Endogenous peroxidase blocking: 3% H₂O₂, 10 minutes at room temperature.
Blocking: 1% BSA + 10% normal goat serum, 10 minutes at room temperature.
Primary antibody: HA752010, 1/50, 1 hour at room temperature.
Secondary antibody: HA1119, 20 minutes at room temperature.
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"