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    dnmt3a-antibody-ab4897.pdf

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Epigenetics and Nuclear Signaling DNA methylation DNMTs
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Anti-Dnmt3a antibody (ab4897)

  • Datasheet
Reviews (4)Q&A (7)References (12)

Product price, shipping and contact information

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Western blot - Anti-Dnmt3a antibody (ab4897)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Dnmt3a antibody (ab4897)

Key features and details

  • Rabbit polyclonal to Dnmt3a
  • Suitable for: IHC-P, WB
  • Reacts with: Human
  • Isotype: IgG

You may also be interested in

Protein
Product image
Recombinant human Dnmt3a protein (ab170408)
Secondary
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Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
Knockout
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Human DNMT3A knockout HeLa cell line (ab261793)

View more associated products

Overview

  • Product name

    Anti-Dnmt3a antibody
    See all Dnmt3a primary antibodies
  • Description

    Rabbit polyclonal to Dnmt3a
  • Host species

    Rabbit
  • Tested Applications & Species

    Application Species
    IHC-P
    Human
    WB
    Human
    See all applications and species data
  • Immunogen

    Synthetic peptide corresponding to Human Dnmt3a aa 457-486 conjugated to Keyhole Limpet Haemocyanin (KLH).

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer

    Preservative: 0.09% Sodium azide
    Constituent: PBS
  • Concentration information loading...
  • Purity

    Ammonium Sulphate Precipitation
  • Purification notes

    This antibody is prepared by Saturated Ammonium Sulfate (SAS) precipitation followed by dialysis against PBS.
  • Clonality

    Polyclonal
  • Isotype

    IgG
  • Research areas

    • Epigenetics and Nuclear Signaling
    • DNA methylation
    • DNMTs

Associated products

  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
  • Isotype control

    • Rabbit IgG, polyclonal - Isotype Control (ChIP Grade) (ab171870)
  • Recombinant Protein

    • Recombinant human Dnmt3a protein (ab170408)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab4897 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Guaranteed

Tested applications are guaranteed to work and covered by our Abpromise guarantee.

Predicted

Predicted to work for this combination of applications and species but not guaranteed.

Incompatible

Does not work for this combination of applications and species.

Application Species
IHC-P
Human
WB
Human
All applications
Mouse
Rat
Chicken
Application Abreviews Notes
IHC-P
1/50 - 1/100.
WB
1/1000. Detects a band of approximately 101 kDa.
Notes
IHC-P
1/50 - 1/100.
WB
1/1000. Detects a band of approximately 101 kDa.

Target

  • Function

    Required for genome wide de novo methylation and is essential for the establishment of DNA methylation patterns during development. DNA methylation is coordinated with methylation of histones. It modifies DNA in a non-processive manner and also methylates non-CpG sites. May preferentially methylate DNA linker between 2 nucleosomal cores and is inhibited by histone H1. Plays a role in paternal and maternal imprinting. Required for methylation of most imprinted loci in germ cells. Acts as a transcriptional corepressor for ZNF238. Can actively repress transcription through the recruitment of HDAC activity.
  • Tissue specificity

    Highly expressed in fetal tissues, skeletal muscle, heart, peripheral blood mononuclear cells, kidney, and at lower levels in placenta, brain, liver, colon, spleen, small intestine and lung.
  • Sequence similarities

    Belongs to the C5-methyltransferase family.
    Contains 1 ADD domain.
    Contains 1 GATA-type zinc finger.
    Contains 1 PHD-type zinc finger.
    Contains 1 PWWP domain.
  • Domain

    The PWWP domain is essential for targeting to pericentric heterochromatin.
  • Post-translational
    modifications

    Sumoylated; sumoylation disrupts the ability to interact with histone deacetylases (HDAC1 and HDAC2) and repress transcription.
  • Cellular localization

    Nucleus. Cytoplasm. Accumulates in the major satellite repeats at pericentric heterochromatin.
  • Target information above from: UniProt accession Q9Y6K1 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 421991 Chicken
    • Entrez Gene: 1788 Human
    • Entrez Gene: 13435 Mouse
    • Entrez Gene: 444984 Rat
    • Omim: 602769 Human
    • SwissProt: Q4W5Z4 Chicken
    • SwissProt: Q9Y6K1 Human
    • SwissProt: O88508 Mouse
    • SwissProt: Q1LZ53 Rat
    • Unigene: 515840 Human
    • Unigene: 5001 Mouse
    • Unigene: 92659 Rat
    see all
  • Alternative names

    • DNA (cytosine 5) methyltransferase 3 alpha antibody
    • DNA (cytosine 5) methyltransferase 3A antibody
    • DNA (cytosine-5)-methyltransferase 3A antibody
    • DNA cytosine methyltransferase 3A2 antibody
    • DNA methyltransferase 3 alpha antibody
    • DNA methyltransferase 3a antibody
    • DNA methyltransferase HsaIIIA antibody
    • DNA MTase HsaIIIA antibody
    • DNM3A_HUMAN antibody
    • DNMT 3a antibody
    • DNMT antibody
    • Dnmt3a antibody
    • DNMT3A2 antibody
    • M.HsaIIIA antibody
    • MCMT antibody
    • OTTHUMP00000201149 antibody
    • TBRS antibody
    see all

Images

  • Western blot - Anti-Dnmt3a antibody (ab4897)
    Western blot - Anti-Dnmt3a antibody (ab4897)
    Anti-Dnmt3a antibody (ab4897) + T47-D cell lysate

    Secondary
    HRP-conjugated anti-rabbit IgG
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Dnmt3a antibody (ab4897)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Dnmt3a antibody (ab4897)

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human hepatocarcinoma tissue labelling Dnmt3a with ab4897.

Protocols

  • Western blot protocols
  • Immunohistochemistry protocols
  • Immunocytochemistry & immunofluorescence protocols

Click here to view the general protocols

Datasheets and documents

    • Datasheet
  • References (12)

    Publishing research using ab4897? Please let us know so that we can cite the reference in this datasheet.

    ab4897 has been referenced in 12 publications.

    • Yuan H  et al. Hypomethylation of nerve growth factor (NGF) promotes binding of C/EBPa and contributes to inflammatory hyperalgesia in rats. J Neuroinflammation 17:34 (2020). PubMed: 31980031
    • Li X  et al. MCM3AP-AS1 KD Inhibits Proliferation, Invasion, and Migration of PCa Cells via DNMT1/DNMT3 (A/B) Methylation-Mediated Upregulation of NPY1R. Mol Ther Nucleic Acids 20:265-278 (2020). PubMed: 32193153
    • Xu X  et al. The long non-coding RNA Linc-GALH promotes hepatocellular carcinoma metastasis via epigenetically regulating Gankyrin. Cell Death Dis 10:86 (2019). PubMed: 30692513
    • Akiyama N  et al. Regulation of DNA methylation levels in the process of oral mucosal regeneration in a rat oral ulcer model. Histol Histopathol N/A:18147 (2019). PubMed: 31286466
    • Ma M  et al. Preeclampsia is associated with hypermethylation of IGF-1 promoter mediated by DNMT1. Am J Transl Res 10:16-39 (2018). IHC-P ; Human . PubMed: 29422991
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-10 of 11 Abreviews or Q&A

    Western blot abreview for Anti-Dnmt3a antibody

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Purified protein (Brain (Cortex))
    Gel Running Conditions
    Reduced Denaturing (8% gel)
    Loading amount
    20 µg
    Specification
    Brain (Cortex)
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 22°C
    Read More

    Diana Christian

    Verified customer

    Submitted May 03 2018

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Dnmt3a antibody

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Sample
    Rat Tissue sections (Testis)
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citric acid
    Permeabilization
    No
    Specification
    Testis
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
    Fixative
    Formaldehyde
    Read More

    Mr. Carl Hobbs

    Verified customer

    Submitted Dec 14 2015

    Question

    IHC-FoFr with mouse tissue
    sees cytoplasmic staining but no nuclear staining
    perm:TX-100
    fix: PFA 30 min
    block: NGS
    Ab: 1/800 - 1/50
    AR: tried low heat
    detection: Cy3, works well

    Read More

    Abcam community

    Verified customer

    Asked on Nov 05 2012

    Answer

    Thank you for your phone call.
    I am sorry that the antibody is not working as expected. As we discussed, I have arranged for a free of charge replacement with ab4897. Please see details below. The item is currently backordered, but the estimated delivery date is set for xxx. I Hope that is OK.

    Also, below please find the discount code for trying ab4897 in mouse and rat and additional information on how it works.

    DISCOUNT CODE: xxx
    Expiration date: xxx
    Value: xxx

    I am very pleased to hear you would like to accept our offer and test ab4897 in mouse and rat. This code will give you xxx off your next order before the expiration date. To redeem this offer, please submit an Abreview for mouse and/or rat and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. Please remember that submission of the Abreview is sufficient for the discount code to become active.
    For more information on how to submit an Abreview, please visit the site: www.abcam.com/Abreviews.

    Remember, we publish both positive and negative Abreviews on our datasheets so please submit the results of your tests. The code will be active once the Abreview has been submitted and can be redeemed in one of the following ways: 1) Call to place your order and mention the code to our customer service department; 2) Include the code in your fax order; 3) Place your order on the web and enter the promotional code.

    Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research.

    The terms and conditions applicable to this offer can be found here: www.abcam.com/abtrial.

    Here is the information regarding your free of charge replacement:

    I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with the order number xxx with ab4897.

    To check the status of the order please contact our Customer Service team and reference this number.

    Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

    I wish you the best of luck with your research.

    Read More

    Abcam Scientific Support

    Answered on Nov 05 2012

    Question

    I have a question regarding the cross-species reactivity of the Anti-Dnmt3a antibody (ab4897). Can you confirm whether the synthetic peptide used to raise the antibody is 100% homolog to rat and mouse? I am also interested if further experimental evidence for the antibodies cross-species reactivity (e.g. western analysis with negative or positive result) exists?

    Read More

    Abcam community

    Verified customer

    Asked on Sep 07 2012

    Answer

    Thank you very much for your interest in ab4897.

    To our knowledge, ab4897 has not been tested in Mouse and Rat.It is very likely that ab4897works in these species since thehomolgy between the immunogen peptide and the proteins of interest is 100% (See http://web.expasy.org/cgi-bin/blast/blast.pl?action=HTML&sequence=DERTRERLVYEVRQK). Therefore, I can offer a discount off a future purchase if you buy ab4897 now, test it in Mouse and Rat and submit feedback to us in the form of an Abreview. It doesn’t matter whether the Abreview is positive or negative, we would just really like to receive your feedback. The discount would be to the value of 1 free primary antibody.

    If you are interested in this offer, please follow these steps:

    1. Reply to this e-mail to let me know that you would like to proceed and test ab4897 in Mouse and Rat. I will then send a discount code. This code must be issued before purchasing ab4897 so please wait for my reply before ordering.

    2. Purchase ab4897 either by phone, fax, or online (https://www.abcam.com). No need to mention the code at this stage.

    3. Test it in Mouse and Rat.

    4. Let us know the results, positive or negative, using our Abreview system (this will take about 10 minutes and images are great if you have them!). To find out how to submit an Abreview, please visit: https://www.abcam.com/abreviews.

    5. After the review is submitted to us, the discount code becomes active. Simply place your new order by phone, fax, or on the web and mention the discount code. The discount can be redeemed for anyprimary antibodyordered and the discount code is valid for 4 months after issue.
    Please remember that submission of the Abreview is sufficient for the discount code to become active.

    We are always pleased to obtain feedback about our products and any information is greatly appreciated! Even if ab4897 turns out to be unsuitable for Mouse and Rat, you will still receive the discount on your next purchase after your Abreview has been submitted.

    Please let me know if you have any questions about this offer and I would be happy to help you further.

    The Terms and Conditions of this offer can be found at: https://www.abcam.com/collaborationdiscount.

    Read More

    Abcam Scientific Support

    Answered on Sep 07 2012

    Question

    Dear Technical Team,
    Thank you for your extensive reply - there's lots for me to try out!
    Please could you tell me if the blocking peptide for ab119282 is
    available to purchase from Abcam and if so could you provide me with a
    product code and price?
    Thanks once again for your help,

    Read More

    Abcam community

    Verified customer

    Asked on Jul 17 2012

    Answer

    Thank you for your reply.
    I am sorry to confirm it is not possible to provide a blocking peptide on this occasion. I apologise for the inconvenience, I understand this would have been useful in this case.

    I hope the other suggestions provided will be helpful and look forward to hearing from you with the outcome. Please do not hesitate to let me know if you have any further concerns regarding the results. I hope we can resolve this case for you as soon as possible.

    Read More

    Abcam Scientific Support

    Answered on Jul 17 2012

    Question

    Inquiry: Antibodies ab4897 lot GR32135-2 and ab119282 lot GR75331-1 (anti Dnmt3a and anti Dnmt3b). I am looking at cytoplasmic and nuclear lysates from HT29 and C4 II cells. I have used an NK-PER kit from Pierce to prepare the lysates, protease inhibitors included. I have run 7.5% SDS PAGE Gels and have blotted onto nitrocellulose membranes. I have used various blocking buffers including TBS/T (+ 5% or 10% Milk Powder), TBS/T + 1%BSA, TBS/T+0.2% fish gelatin, blocking with each at RT and overnight at 4C. I have varied my concentrations of both primary and secondary Abs. I have varied my incubation times for primary Abs and incubation temperatures, over night at 4C or 1 hr to 3hrs at RT. I have used an Anti GAPDH and Anti Dnmt1 both purchased from Abcam at the same time as the Dnmt3a and 3b and have had no problem with those whatsoever. However, with the Anti-Dnmt3a and Anti Dnmt3b I consistantly get multiple strong bands, just about everywhere except at the required molecular weights (I have had faint positivity for Dnmt3b in a HT29 nuclear extract along with multiple other bands). The fact that the Dnmt1 and GAPDH antibodies seem to work very well suggests that the electrophoresis and transfer conditions are OK and the ECL kit is fine. Please could you offer advice as I'm just about running out of ideas? Please could you tell me if there is some information about the nature of the antibodies that you think I am failing to realise? I am using HT29 as a control for Dnmt3b as suggested on your data sheet and our cell line has been verified fairly recently. Is it possible for you to send me an outline of the protocol used to produce the image on the data sheet as my images are not at all similar? Thank you for your help,

    Read More

    Abcam community

    Verified customer

    Asked on Jul 11 2012

    Answer

    Thank you for taking the time to contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from these antibody.

    I would like to reassure you that these antibodies are tested and covered by our 6 month guarantee for WB and human samples. In the event that a product is not functioning in the tested applications and species cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a creditnote, free of charge replacement or refund.

    As requested, I am forwarding a copy of the general WB testing protocols for these antibodies. Please note that these will be a guideline only and may require some further individual optimization by the end user.

    ab4897 Anti-Dnmt3a antibody:
    A. Preparation of cell lysates
    1. Collect cells (confluent T-25) by trypsinization and spin.
    2. Lyze the pellet with 100 ul lysis buffer on ice for 10 min. For 500,000 cells, lyze with 20 ul.
    3. Spin at 14,000 rpm (16,000 g) in an Eppendorf microfuge for 10 min at 4‹C.
    4. Transfer the supernatant to a new tube and discard the pellet.
    5. Determine the protein concentration (Bradford assay, A280, or BCA)
    6. Take x ul (= y ug protein) and mix with x ul of 2x sample buffer.
    7. Boil for 5 min and cool at RT for 5 min.
    8. Flash spin to bring down condensation prior to loading gel.
    B. Polyacrylamide gel (14.5 cm ˜ 16.5 cm)
    1. Agarose plug:
    1% agarose dissolved in 1 ˜ Resolving gel buffer.
    2. Resolving gel:
    24 ml of a 9% gel
    5.4 ml 40% acrylamide/bisacrylamide (29:1 mix)
    3 ml 8x Resolving gel buffer
    15.6 ml water
    12 ul TEMED
    60 ul 20% ammonium persulfate
    3. Stacking gel: 8 ml
    1 ml 40% acrylamide/bisacrylamide (29:1 mix)
    2 ml 4 ˜ Stacking gel buffer
    5 ml water
    8 ul TEMED
    21.6 ul 20% ammonium persulfate
    C. Preparation of gel
    1. Assemble the glass plates and spacers (1.5 mm thick).
    2. Pour an agarose plug (1-2 mm).
    3. Pour the running gel to about 1 cm below the wells of the comb (˜20 ml).
    4. Seal with 1 ml water-saturated 1-butanol. (Can stop here and leave gel as is overnight if you want.)
    5. When gel has set, pour off the butanol and rinse with deionized water.
    6. Pour the stacking gel (˜5 ml) and insert the comb immediately.
    7. When the stacking gel has set, place in gel rig and immerse in buffer.
    8. Prior to running the gel, flush the wells out thoroughly with running buffer.
    D. Running the gel
    1. After flash spinning the samples, load into the wells.
    2. Be sure to use markers. We use 15 ul Bio-Rad Kaleidoscope Prestained Standards #161-0324 directly.
    3. Run with constant current (35-37 mA with voltage set at >150 V).
    4. Usual running time is about 1.3 hr.
    E. Using precast gels (Ready Gels from Bio-Rad)
    1. Assemble gel in gel rig.
    2. Prepare protein samples (10 ug will suffice).
    3. Use 5 ul of Kaleidoscope standard.
    4. Run at 200 V (constant voltage) for 30 min.
    F. Preparation of membrane
    1. Cut a piece of PVDF membrane.
    2. Wet in methanol on a rocker at RT for 5 mins. Remove methanol and add 1x Transfer buffer until ready to use.
    G. Membrane transfer
    1. Assemble "sandwich":
    2. Prewet the sponges, filter papers (slightly bigger than gel) in 1 ˜ Blotting buffer. Sponge - filter paper - gel - membrane - filter paper - sponge
    3. Transfer for 1 hr at 15 volts at 4oC on a stir plate. Bigger proteins might take longer to transfer. For a Mini-Transblot, it's 100 V for 1 hr with the cold pack and prechilled buffer.
    4. Immerse membrane in Amido-Black stain 5 mins.
    5. Destain 4 ˜ 5 mins with destaining buffer.
    6. When finished, immerse membrane in Blocking buffer and block for one hour at room temperature.
    H. Antibodies and detection
    1. Incubate with primary antibody diluted to 2 ug/ml in total volume of 3 mL in Blocking buffer for one hour at room temperature.
    2. Wash 4 ˜ 5 min with 0.05% Tween 20 in TBS.
    3. Incubate with secondary antibody diluted 1:10,000 (eg HRP anti-rabbit) in Blocking buffer for 1 hour at room temp.
    4. Wash 4 ˜ 5 min with 0.05% Tween 20 in TBS.
    5. Detect with Chemiluminescent kit.
    I. Stripping blot
    1. Rinse blot off with 0.05% Tween 20 in PBS.
    2. Put blot into Kapak bag cut to slightly bigger size than blot.
    3. Add about 5 to 10 ml Stripping buffer.
    4. Remove as much air as possible and seal bag.
    5. Immerse into 80‹C water bath and incubate for 20 min.
    6. Rinse blot off with 0.05% Tween 20 in PBS.
    7. Block for about 1 hr with 5% BSA/Tween 20, or overnight with 3% BSA/Tween 20.
    Buffers for Westerns
    Lysis buffer:
    0.15 M NaCl
    5 mM EDTA, pH 8
    1% Triton X100
    10 mM Tris-Cl, pH 7.4
    Just before using add:
    1:1000 5 M DTT
    1:1000 100 mM PMSF in isopropanol
    1:1000 5 M ƒÃ.aminocaproic acid
    2x sample buffer:
    130 mM Tris-Cl, pH8.0
    20% (v/v) Glycerol
    4.6% (w/v) SDS
    0.02% Bromophenol blue
    2% DTT
    8x Resolving gel buffer: 100 ml
    0.8 g SDS (add last)
    36.3 g Trizma base (= 3 M)
    Adjust pH to 8.8 with concentrated HCl
    4x Stacking gel buffer: 100 ml
    0.4 g SDS (add last)
    6.05 g Trizma base (= 0.5 M)
    Adjust pH to 6.8
    10x Running buffer: 1 L
    30.3 g Trizma base (= 0.25 M)
    144 g Glycine (= 1.92 M)
    10 g SDS (= 1%)--add last


    ab119282 Anti-Dnmt3b antibody:
    Western blot Protocol (Colorimetric)
    Blocking
    1. Block membrane by incubating 1 hour at room temperature with shaking in Blocking Solution (5% nonfat dry milk, 0.05% Tween-20 in TBS (50mM Tris, 100mM NaCl, pH 7.6).
    Incubation with Primary Antibody
    1. Dilute primary antibody at the appropriate dilution in Blocking Solution.
    2. Incubate the membrane with diluted primary antibody for overnight at 4℃ with agitation.
    3. Remove antibody solution. Wash the membrane 3 times for 10 minutes each time at room temperature in TBST (50mM Tris, 100mM NaCl, 0.05% Tween-20, pH 7.6) with shaking. Note: Increase the concentration of Tween-20 to 0.1% reduces the background and increases the specificity, but it will reduce the sensitivity.
    Incubation with Second Antibody
    1. Incubate membrane with secondary AP conjugate diluted (according to manufacturer’s instructions) in Blocking Solution for 1 hour at room temperature with shaking.
    2. Wash the membrane 3 times for 10 minutes each time at room temperature in TBST (50mM Tris, 100mM NaCl, 0.05% Tween-20, pH 7.6) with shaking.
    3. Wash membrane with TBS for 2-5 minutes.
    Stain
    1. Warm up BCIP/NBT Solution (according to manufacturer’s instructions) to room temperature before use.
    2. Apply enough of BCIP/NBT solution to cover the specimen completely. Allow 5-10 minutes, under room temperature, for the color to develop.
    3. Monitor the color development until desired intensity is observed.
    4. Rinse the specimen gently with distilled water to stop the reaction.
    Peptide Competition
    Before proceeding Western Immunoblotting, add Blocking Peptide to the diluted primary antibody in a molar ratio of 10:1 (peptide to antibody) and incubate the mixture at 4℃ for overnight or at room temperature for 2 hours.
    For ab119282, 50 ug of total protein (HT29 cell lysates) was loaded in each lane on an 10% SDS-Page gel. The antibody was used at a concentration of 1 ug/ml diluted in 1% nonfat dry milk, 0.05% Tween-20 in TBS.

    I hope these will be helpful to you. However, if you still have some concerns regarding the results I would be pleased to investigate this particular case further for you, and also obtain more information for our quality monitoring records. In order to proceed with this, I have enclosed a technical questionnaire below. I would appreciate if you could complete this. It will help you put the information we require together very easily.

    I would appreciate if you are also able to provide images which would help us to assess the results.

    Thank you for your time and cooperation. We look forward to receiving the completed questionaire.

    Order Details
    Antibody code:
    Problem
    Choose: Non-specific band Multiple bands No signal or weak signal High background
    Lot number
    Purchase order number
    or preferably Abcam order number:
    General Information
    Antibody storage conditions (temperature/reconstitution etc)
    Description of the problem (high background, wrong band size, more bands, no band etc.)
    Sample (Species/Cell extract/Nuclear extract/Purified protein/Recombinant protein etc.)
    Sample preparation (Buffer/Protease inhibitors/Heating sample etc.)
    Amount of protein loaded

    Electrophoresis/Gel conditions (Reducing or Non-reducing gel, % of the gel etc.)
    Transfer and blocking conditions (Buffer/time period, Blocking agent etc.)
    Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)
    Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)
    Detection method (ECL, ECLPlus etc.)
    Positive and negative controls used (please specify)
    Optimization attempts (problem solving)
    How many times have you tried the Western?
    Have you run a "No Primary" control?
    Yes No
    Do you obtain the same results every time?
    Yes No
    e.g. are the background bands always in the same place?
    What steps have you altered?
    Additional Notes:
    Image:
    We would appreciate if you are able to provide an image (including molecular weight markers) which would help us to asess the results.

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    Abcam Scientific Support

    Answered on Jul 11 2012

    Immunoprecipitation abreview for Anti-Dnmt3a antibody

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunoprecipitation
    Sample
    Human Cell lysate - whole cell (lung carcinoma)
    Total protein in input
    300 µg
    Specification
    lung carcinoma
    Immuno-precipitation step
    Protein G
    Read More

    Abcam user community

    Verified customer

    Submitted Jan 19 2011

    Immunocytochemistry/ Immunofluorescence abreview for Anti-Dnmt3a antibody

    Good
    Abreviews
    Abreviews
    abreview image
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (hESC, mESC)
    Specification
    hESC, mESC
    Fixative
    Paraformaldehyde
    Permeabilization
    Yes - 0.1% Triton X-100 in PBS for 15min at room temp.
    Blocking step
    BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 4% · Temperature: 21°C
    Read More

    Abcam user community

    Verified customer

    Submitted Sep 11 2007

    Question

    What is recommended to use as a positive control for this antibody in Western blotting?

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    Abcam community

    Verified customer

    Asked on Oct 14 2004

    Answer

    The originator used T47-D cell lysate as a positive control for ab4897 in Western blotting. If you have any more questions, please let us know.

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    Abcam Scientific Support

    Answered on Oct 15 2004

    Question

    I want to know if this Dnmt3a antibody (ab4897) could work on IHC-P Immunohistochemistry (Formalin-fixed paraffin-embedded sections)

    Read More

    Abcam community

    Verified customer

    Asked on Nov 09 2003

    Answer

    To our knowledge, this antibody has yet to be tested in this application. All tested applications are specified on Abcam product datasheets. If you decide to go ahead and purchase this product, please let us know how you get on and in return we will forward a reward of your choice, typically an Amazon gift voucher.

    Read More

    Abcam Scientific Support

    Answered on Nov 10 2003

    1-10 of 11 Abreviews or Q&A

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