MBD1 Antibody (100B272.1) - BSA Free

Novus Biologicals | Catalog # NB100-56537

Novus Biologicals
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Key Product Details

Validated by

Knockout/Knockdown

Species Reactivity

Validated:

Human, Mouse

Cited:

Human, Mouse

Applications

Validated:

Immunohistochemistry, Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, Chromatin Immunoprecipitation, Chromatin Immunoprecipitation (ChIP), Knockdown Validated

Cited:

Immunohistochemistry-Frozen, Western Blot, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, Chemotaxis

Label

Unconjugated

Antibody Source

Monoclonal Mouse IgG1 Clone # 100B272.1

Format

BSA Free
View all MBD1 Primary Antibodies »

Product Specifications

Immunogen

This antibody was generated by immunizing mice with a synthetic peptide corresponding to amino acids 391-405 (SESEDGAGSPPPYRR) of human MBD1; GenBank no ref|NP_056671.2|

Reactivity Notes

Mouse reactivity reported in scientific literature (PMID: 25798578)

Specificity

There are many isoforms of human MBD1 ranging from 503 (~55 kDa) to 655 (~71 kDa) amino acids in length. Researchers are encouraged to use the provided immunogen sequence to determine if this antibody is suitable for testing a specific isoform of interest.

Clonality

Monoclonal

Host

Mouse

Isotype

IgG1

Scientific Data Images for MBD1 Antibody (100B272.1) - BSA Free

Knockdown Validated: MBD1 Antibody (100B272.1) [NB100-56537]

Western Blot: MBD1 Antibody (100B272.1) [NB100-56537]

Western Blot: MBD1 Antibody (100B272.1) [NB100-56537] - blot shows lysates of HEK293T human embryonic kidney parental cell line and MBD1 knockout (KO) HEK293T cell line. PVDF membrane was probed with 2.0 ug/ml of Mouse Anti-Human MBD1 Monoclonal Antibody (Catalog # NB100-56537) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog #HAF018). Specific band was detected for MBD1 at approximately 60 kDa (as indicated) in the parental HEK293T cell line, but is not detectable in the knockout HEK293T cell line. This experiment was conducted under reducing conditions.
Western Blot: MBD1 Antibody (100B272.1) [NB100-56537]

Western Blot: MBD1 Antibody (100B272.1) [NB100-56537]

Western Blot: MBD1 Antibody (100B272.1) [NB100-56537] - Analysis using Azide and BSA Free version of NB100-56537. Jurkat cell lysate using anti-MBD1 antibody. Image from verified customer review.
Immunocytochemistry/ Immunofluorescence: MBD1 Antibody (100B272.1) [NB100-56537]

Immunocytochemistry/ Immunofluorescence: MBD1 Antibody (100B272.1) [NB100-56537]

Immunocytochemistry/Immunofluorescence: MBD1 Antibody (100B272.1) [NB100-56537] - NIH3T3 cells were fixed for 10 minutes using 4% PFA and then permeabilized for 5 minutes using 1X PBS + 0.5% Triton-X100. The cells were incubated with anti-MBD1 (100B272.1) at 2 ug/ml overnight at 4C and detected with an anti-mouse Dylight 488 (Green) at a 1:500 dilution. Actin was detected with Phalloidin 568 (Red) at a 1:500 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.
Immunohistochemistry-Paraffin: MBD1 Antibody (100B272.1) [NB100-56537]

Immunohistochemistry-Paraffin: MBD1 Antibody (100B272.1) [NB100-56537]

Immunohistochemistry-Paraffin: MBD1 Antibody (100B272.1) [NB100-56537] - Analysis of a FFPE tissue section of human colon using 1:200 dilution of MBD1 (100B272.1) antibody. The staining was developed using HRP labeled anti-mouse secondary antibody and DAB reagent, and nuclei of cells were counter-stained with hematoxylin. Cytoplasmic and nuclear staining was observed.
Western Blot: MBD1 Antibody (100B272.1) [NB100-56537]

Western Blot: MBD1 Antibody (100B272.1) [NB100-56537]

Western Blot: MBD1 Antibody (100B272.1) [NB100-56537] - Analysis of MBD1 in HeLa lysate using MBD1 antibody at 2 ug/ml (lane 1) and 1 ug/ml (lane 2).
Immunocytochemistry/ Immunofluorescence: MBD1 Antibody (100B272.1) [NB100-56537]

Immunocytochemistry/ Immunofluorescence: MBD1 Antibody (100B272.1) [NB100-56537]

Immunocytochemistry/Immunofluorescence: MBD1 Antibody (100B272.1) [NB100-56537] - HeLa cells were fixed for 10 minutes using 4% PFA and then permeabilized for 5 minutes using 1X PBS + 0.5% Triton-X100. The cells were incubated with anti-MBD1 (100B272.1) at 2 ug/ml overnight at 4C and detected with an anti-mouse Dylight 488 (Green) at a 1:500 dilution. Actin was detected with Phalloidin 568 (Red) at a 1:500 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.

Applications for MBD1 Antibody (100B272.1) - BSA Free

Application
Recommended Usage

Chromatin Immunoprecipitation

1:10-1:500. Use reported in scientific literature (PMID 25798578)

Chromatin Immunoprecipitation (ChIP)

1:10-1:500

Immunocytochemistry/ Immunofluorescence

reported in scientific literature (PMID 24464130)

Immunohistochemistry

1:200

Immunohistochemistry-Frozen

reported in scientific literature (PMID 24464130)

Immunohistochemistry-Paraffin

1:200

Immunoprecipitation

1:10-1:500. Use reported in scientific literature (PMID 15327775)

Western Blot

2-4 ug/ml

Reviewed Applications

Read 2 reviews rated 4 using NB100-56537 in the following applications:

Formulation, Preparation, and Storage

Purification

Protein G purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: MBD1

DNA methylation, or the addition of methyl groups to cytosine bases in the dinucleotide CpG, is imperative to proper development and regulates gene expression. The methylation pattern involves the enzymatic processes of methylation and demethylation. The demethylation enzyme was recently found to be a mammalian protein, which exhibits demethylase activity associated to a methyl-CpG-binding domain (MBD). The enzyme is able to revert methylated cytosine bases to cytosines within the particular dinucleotide sequence mdCpdG by catalyzing the cleaving of the methyl group as methanol. MeCP2 and MBD1 (PCM1) are first found to repress transcription by binding specifically to methylated DNA. MBD2 and MBD4 (also known as MED1) were later found to colocalize with foci of heavily methylated satellite DNA and believed to mediate the biological functions of the methylation signal. Surprisingly, MBD3 does not bind methylated DNA both in vivo and in vitro. MBD1, MBD2, MBD3, and MBD4 are found to be expressed in somatic tissues, but the expression of MBD1 and MBD2 is reduced or absent in embryonic stem cells, which are known to be deficient in MeCP1 activity. MBD4 have homology to bacterial base excision repair DNA N-glycosylases/lyases. In some microsatellite unstable tumors MBD4 is mutated at an exonic polynucleotide tract.

Alternate Names

CXXC-type zinc finger protein 3, methyl-CpG binding domain protein 1, methyl-CpG binding domain protein 1 isoform PCM1, methyl-CpG-binding domain protein 1, Methyl-CpG-binding protein MBD1, PCM1CXXC3RFT, Protein containing methyl-CpG-binding domain 1, the regulator of fibroblast growth factor 2 (FGF-2) transcription

Gene Symbol

MBD1

UniProt

Q9UIS9 (Human)

Additional MBD1 Products

Product Documents for MBD1 Antibody (100B272.1) - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

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Product Specific Notices for MBD1 Antibody (100B272.1) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Citations for MBD1 Antibody (100B272.1) - BSA Free

Customer Reviews for MBD1 Antibody (100B272.1) - BSA Free (2)

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  • MBD1 Antibody (100B272.1)
    Name: Anonymous
    Application: Western Blot
    Sample Tested: cell lysate
    Species: Mouse
    Verified Customer | Posted 08/06/2021
    Western blot, cell lysates.
    MBD1 Antibody (100B272.1) - BSA Free NB100-56537
  • Name: Anonymous
    Application: Western Blot
    Sample Tested: Sample tested: Jurkat cells
    Species: Human
    Verified Customer | Posted 10/07/2014
    Western for MBD1 in Jurkat cells
    MBD1 Antibody (100B272.1) - BSA Free NB100-56537

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Protocols

View specific protocols for MBD1 Antibody (100B272.1) - BSA Free (NB100-56537):

Immunocytochemistry Protocol

Culture cells to appropriate density in 35 mm culture dishes or 6-well plates.

1. Remove culture medium and wash the cells briefly in PBS. Add 10% formalin to the dish and fix at room temperature for 10 minutes.
2. Remove the formalin and wash the cells in PBS.
3. Permeablize the cells with 0.1% Triton X100 or other suitable detergent for 10 min.
4. Remove the permeablization buffer and wash three times for 10 minutes each in PBS. Be sure to not let the specimen dry out.
5. To block nonspecific antibody binding, incubate in 10% normal goat serum from 1 hour to overnight at room temperature.
6. Add primary antibody at appropriate dilution and incubate overnight at 4C.
7. Remove primary antibody and replace with PBS. Wash three times for 10 minutes each.
8. Add secondary antibody at appropriate dilution. Incubate for 1 hour at room temperature.
9. Remove secondary antibody and replace with PBS. Wash three times for 10 minutes each.
10. Counter stain DNA with DAPi if required.

Immunohistochemistry-Paraffin Embedded Sections

Antigen Unmasking:
Bring slides to a boil in 10 mM sodium citrate buffer (pH 6.0) then maintain at a sub-boiling temperature for 10 minutes. Cool slides on bench-top for 30 minutes (keep slides in the sodium citrate buffer all the time).

Staining:
1. Wash sections in deionized water three times for 5 minutes each.
2. Wash sections in PBS for 5 minutes.
3. Block each section with 100-400 ul blocking solution (1% BSA in PBS) for 1 hour at room temperature.
4. Remove blocking solution and add 100-400 ul diluted primary antibody. Incubate overnight at 4 C.
5. Remove antibody solution and wash sections in wash buffer three times for 5 minutes each.
6. Add 100-400 ul HRP polymer conjugated secondary antibody. Incubate 30 minutes at room temperature.
7. Wash sections three times in wash buffer for 5 minutes each.
8. Add 100-400 ul DAB substrate to each section and monitor staining closely.
9. As soon as the sections develop, immerse slides in deionized water.
10. Counterstain sections in hematoxylin.
11. Wash sections in deionized water two times for 5 minutes each.
12. Dehydrate sections.
13. Mount coverslips.


1. Perform SDS-PAGE on samples to be analyzed, loading 10-25 ug of total protein per lane.
2. Transfer proteins to PVDF membrane according to the instructions provided by the manufacturer of the membrane and transfer apparatus.
3. Stain the membrane with Ponceau S (or similar product) to assess transfer success, and mark molecular weight standards where appropriate.
4. Rinse the blot TBS -0.05% Tween 20 (TBST).
5. Block the membrane in 5% Non-fat milk in TBST (blocking buffer) for at least 1 hour.
6. Wash the membrane in TBST three times for 10 minutes each.
7. Dilute primary antibody in blocking buffer and incubate overnight at 4C with gentle rocking.
8. Wash the membrane in TBST three times for 10 minutes each.
9. Incubate the membrane in diluted HRP conjugated secondary antibody in blocking buffer (as per manufacturer's instructions) for 1 hour at room temperature.
10. Wash the blot in TBST three times for 10 minutes each (this step can be repeated as required to reduce background).
11. Apply the detection reagent of choice in accordance with the manufacturers instructions.

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