Overview

  • Product name
    Anti-Drosha antibody - ChIP Grade
    See all Drosha primary antibodies
  • Description
    Rabbit polyclonal to Drosha - ChIP Grade
  • Host species
    Rabbit
  • Tested applications
    Suitable for: ICC/IF, IP, WB, ICC, IHC-P, ChIPmore details
  • Species reactivity
    Reacts with: Mouse, Human
  • Immunogen

    Synthetic peptide derived from within residues 1 - 100 of Human Drosha.

    Read Abcam's proprietary immunogen policy (Peptide available as ab12307.)

  • Positive control
    • HeLa cells
  • General notes
    Three isoforms of 159, 156 and 143 kDa.

Properties

Applications

Our Abpromise guarantee covers the use of ab12286 in the following tested applications.

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

Application Abreviews Notes
ICC/IF Use a concentration of 1 µg/ml.
IP Use at an assay dependent concentration.
WB Use a concentration of 1 µg/ml. Predicted molecular weight: 159 kDa.
ICC Use at an assay dependent concentration.
IHC-P 1/100.
ChIP Use at an assay dependent concentration.

Target

  • Function
    Ribonuclease III double-stranded (ds) RNA-specific endoribonuclease that is involved in the initial step of microRNA (miRNA) biogenesis. Component of the microprocessor complex that is required to process primary miRNA transcripts (pri-miRNAs) to release precursor miRNA (pre-miRNA) in the nucleus. Within the microprocessor complex, DROSHA cleaves the 3' and 5' strands of a stem-loop in pri-miRNAs (processing center 11 bp from the dsRNA-ssRNA junction) to release hairpin-shaped pre-miRNAs that are subsequently cut by the cytoplasmic DICER to generate mature miRNAs. Involved also in pre-rRNA processing. Cleaves double-strand RNA and does not cleave single-strand RNA. Involved in the formation of GW bodies.
  • Tissue specificity
    Ubiquitous.
  • Sequence similarities
    Contains 1 DRBM (double-stranded RNA-binding) domain.
    Contains 2 RNase III domains.
  • Domain
    The 2 RNase III domains form an intramolecular dimer where the domain 1 cuts the 3'strand while the domain 2 cleaves the 5'strand of pri-miRNAs, independently of each other.
  • Cellular localization
    Nucleus. Nucleus > nucleolus. A fraction is translocated to the nucleolus during the S phase of the cell cycle. Localized in GW bodies (GWBs), also known as P-bodies.
  • Information by UniProt
  • Database links
  • Alternative names
    • DROSHA antibody
    • Drosha double stranded RNA specific endoribonuclease antibody
    • Drosha ribonuclease type III antibody
    • Etohi2 antibody
    • HSA242976 antibody
    • Nuclear RNase III Drosha antibody
    • p241 antibody
    • Protein Drosha antibody
    • Putative protein p241 which interacts with transcription factor Sp1 antibody
    • Putative ribonuclease III antibody
    • RANSE3L antibody
    • Ribonuclease 3 antibody
    • Ribonuclease III antibody
    • Ribonuclease III nuclear antibody
    • Ribonuclease type III nuclear antibody
    • RibonucleaseIII antibody
    • RN 3 antibody
    • RN3 antibody
    • RNase 3 antibody
    • RNase III antibody
    • RNase3 antibody
    • RNASE3L antibody
    • RNaseIII antibody
    • RNASEN antibody
    • RNC_HUMAN antibody
    see all

Images

  • Human female amniocytes immunostained with ab12286 Drosha (FITC) (1/25 dilution). The DNA is labelled red with propidium iodide. This image was submitted as part of a review by Ahmad Khalil.
  • All lanes : Anti-Drosha antibody - ChIP Grade (ab12286) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat polyclonal to Rabbit IgG (Alexa Fluor® 680) at 1/15000 dilution

    Performed under reducing conditions.

    Predicted band size: 159 kDa
    Observed band size: 170 kDa
    why is the actual band size different from the predicted?

  • ab12286 at a 1/25 dilution staining human HeLa cells by immunocytochemistry. The antibody was incubated with the cells for 1 hour and then detected using a Cy3 conjugated donkey anti-rabbit polyclonal antibody.

    This image is courtesy of an Abreview submitted on 7 February 2006.

    See Abreview

  • All lanes : Anti-Drosha antibody - ChIP Grade (ab12286) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate
    Lane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Nuclear Lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 159 kDa
    Observed band size: 170 kDa why is the actual band size different from the predicted?
    Additional bands at: 115 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 12 minutes


    Abcam recommends using milk as the blocking agent. Abcam welcomes customer feedback and would appreciate any comments regarding this product and the data presented above.
  • ICC/IF image of ab12286 stained HepG2 cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab12286, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
  • ICC/IF image of ab12286 stained Hela cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab12886, 5µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

References

This product has been referenced in:
  • Burger K & Gullerova M Nuclear re-localization of Dicer in primary mouse embryonic fibroblast nuclei following DNA damage. PLoS Genet 14:e1007151 (2018). Read more (PubMed: 29394246) »
  • Hoffmann N  et al. DGCR8 Promotes Neural Progenitor Expansion and Represses Neurogenesis in the Mouse Embryonic Neocortex. Front Neurosci 12:281 (2018). Read more (PubMed: 29760646) »
See all 65 Publications for this product

Customer reviews and Q&As

1-10 of 20 Abreviews or Q&A

Application
Immunoprecipitation
Immuno-precipitation step
Protein G
Sample
Human Cell lysate - nuclear (A549)
Specification
A549
Total protein in input
25 µg

Abcam user community

Verified customer

Submitted Feb 20 2015

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Loading amount
25 µg
Gel Running Conditions
Reduced Denaturing (13.5%)
Sample
Mouse Cell lysate - whole cell (Mouse N2A neuroblastoma)
Specification
Mouse N2A neuroblastoma
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted May 27 2013

Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing (6%)
Sample
Human Cell lysate - whole cell (human osteosarcoma cell line U2OS)
Specification
human osteosarcoma cell line U2OS
Blocking step
Milk as blocking agent for 18 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Abcam user community

Verified customer

Submitted May 21 2013

Question

I performed a western blot with this antibody twice with two different conditions. Both were using protein isolated from three mouse embryonic cell lines (two un-differentiated, one derived into motor neurons). 25ug/lane were loaded for the undifferentiated mESCs and 40ug for the mESC-dervied motor neuron samples (for both westerns). First western: Blocked with 5% milk 30min. Added anti-Drosha antibody 1:1000 (3ug/3ml) in 5% milk. incubated o/n at 4degC on shaker. Washed 3x with PBS+0.5%tween. Added secondary antibody (anti-Rabbit IgG HRP conjugated from donkey, other company) at 1:6000 in 5%milk. Incubated at RT for 1 hour. Washed 3x with PBS+0.5%tween, 2x in PBS. Developed using ECL method. Even after 10 minutes of developing, I could not see any bands whatsoever.

Second western: Blocked with 0.5%milk 30min. Added anti-Drosha antibody 1:500 (6ug/3ml) in PBS. incubated 4 hours RT on shaker. Washed 3x with PBS. Added secondary antibody (anti-Rabbit IgG HRP conjugated from donkey, from other company) at 1:6000 in PBS. Incubated at RT for 1 hour. Washed 3x with PBS. Developed using ECL method. Saw multiple bands at 62kDa, 100kDa, doublet at >250kDa in the two un-differentiated mouse ESC lines. In the differentiated line, I saw bands at 90kDa, 150kDa, 200kDa. Some other faint bands were observed in the lanes. However, there was no band at 150 or 170kDa (the predicted and observed sizes of Drosha) across all three of my samples.
These results suggest that I am not detecting Drosha. I would expect a band at 150kDa (or 170kDa) across all of my protein samples (all are isolated from mouse cells). However, there are many strong bands at other sizes. I would like a refund/credit, since this antibody was recommended for use on mouse and I do not have consistent results in mouse.

Read More
Answer


I can offer you either a free of charge replacement with another Drosha-antibody (e.g. ab135956) or an antibody to a different target. Alternatively, a credit or refund can be arranged.

Read More

Answer

Thank you for your reply.
As requested, I am sending you ab38692 as a free of charge replacement for ab12286 that did not work for you in western blotting. Your new order number is *.
Please let me know if you experience any issues with the new antibody, or if there is anything else I can help you with.
I hope you are having a great day and also that your experiments co-operate.

Read More

Answer

Thank you for your reply.
The antibody, ab12286 is covered under our Abpromise for six months and is guaranteed to work in western blot on rabbit samples . Since we cannot resolve this issue you are having with the antibody then I would be happy to either send a replacement. Please let me know the catalogue number of the antibody you would like to receive as a replacement and I will have it sent out as soon as I can.
I look forward to your reply and helping resolve this issue.

Read More

Answer

Thank you and your customer for your inquiry.
I checked the homology between the immunogen sequence of ab12286 and drosophila and unfortunately have to confirm that there is no significant homology.
Therefore I cannot recommend ab12286 for experiments with drosophila.
Unfortunately, the other two anti Drosha antibodies (ab58589 and ab85027) we have in our catalog at the moment also do not show any promise of a cross reaction with drosophila.
The homology is lower than 45% in all cases.
I am sorry I did not have better news on this occasion and hope this information is nevertheless helpful for your customer.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Mouse Brain Day 25)
Specification
Mouse Brain Day 25
Fixative
neutral buffered formalin
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Sodium Citrate
Permeabilization
Yes - TBS-Tween (0.1%)
Blocking step
Serum as blocking agent for 12 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 4°C

Dr. Kai Orton

Verified customer

Submitted May 28 2012

Answer

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

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 ab58589. Please see below for the discount code for testing this antibody in IP.

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 (for WB, human). Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.


DISCOUNT CODE: xxx
Expiration date: xxx

I am very pleased to hear you would like to accept our offer and test ab58589 in IP. This code will give you: 1 free PRIMARY ANTIBODY before the expiration date. To redeem this offer, please submit an Abreview for IPand include this code in the “Additional Comments” section so we know the Abreview is for this promotion. 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/collaborationdiscount.

I wish you the best of luck with your research.

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1-10 of 20 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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