Anti-BACE1 抗体 (ab2077)

Reviews (4)Q&A (11)References (51)

製品の概要

  • 製品名

    Anti-BACE1 antibody
    BACE1 一次抗体 製品一覧

  • 製品の詳細

    Rabbit polyclonal to BACE1

  • 由来種

    Rabbit

  • アプリケーション

    適用あり: WB, ICC/IF, IHC-Pmore details

  • 種交差性

    交差種: Mouse, Human
    交差が予測される動物種: Rat

  • 免疫原

    Synthetic peptide corresponding to Human BACE1 (C terminal). The immunogen is located within the last 50 amino acids of BACE1 and it consists of a 17 aa peptide.
    Database link: P56817
    (Peptide available as ab7883)

  • ポジティブ･コントロール

    • IHC-P: Mouse brain tissue WB: A431, A549, Caco-2, Daudi, HeLa, K562, Jurkat, SK-N-SH, THP-1. Human brain tissue lysate, 3T3 lysate. ICC/IF: 3T3 cells.

  • 特記事項

    Beta-site APP Cleaving Enzyme.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab2077 in the following tested applications.

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

アプリケーション Abreviews 特記事項
WB Use a concentration of 1 µg/ml. Detects a band of approximately 70 kDa. We recommend overnight incubation at 4C and using 5% skim milk to block.
ICC/IF Use a concentration of 10 µg/ml.
IHC-P Use a concentration of 2.5 µg/ml.

ターゲット情報

  • 機能

    Responsible for the proteolytic processing of the amyloid precursor protein (APP). Cleaves at the N-terminus of the A-beta peptide sequence, between residues 671 and 672 of APP, leads to the generation and extracellular release of beta-cleaved soluble APP, and a corresponding cell-associated C-terminal fragment which is later released by gamma-secretase.

  • 組織特異性

    Expressed at high levels in the brain and pancreas. In the brain, expression is highest in the substantia nigra, locus coruleus and medulla oblongata.

  • 配列類似性

    Belongs to the peptidase A1 family.

  • ドメイン

    The transmembrane domain is necessary for its activity. It determines its late Golgi localization and access to its substrate, APP.

  • 翻訳後修飾

    Glycosylated.

  • 細胞内局在

    Membrane. Golgi apparatus > trans-Golgi network. Endoplasmic reticulum. Endosome. Cell surface. Predominantly localized to the later Golgi/trans-Golgi network (TGN) and minimally detectable in the early Golgi compartments. A small portion is also found in the endoplasmic reticulum, endosomes and on the cell surface.
  • Information by UniProt

  • 参照データベース

    see all

  • 別名

    • APP beta secretase antibody
    • Asp 2 antibody
    • ASP2 antibody
    • Aspartyl protease 2 antibody
    • BACE 1 antibody
    • BACE antibody
    • BACE1 antibody
    • BACE1_HUMAN antibody
    • Beta secretase 1 antibody
    • Beta secretase antibody
    • Beta site amyloid beta A4 precursor protein cleaving enzyme antibody
    • Beta site amyloid precursor protein cleaving enzyme 1 antibody
    • Beta site amyloid precursor protein cleaving enzyme antibody
    • Beta site APP cleaving enzyme 1 antibody
    • Beta site APP cleaving enzyme antibody
    • Beta-secretase 1 antibody
    • Beta-site amyloid precursor protein cleaving enzyme 1 antibody
    • Beta-site APP cleaving enzyme 1 antibody
    • FLJ90568 antibody
    • HSPC104 antibody
    • Memapsin 2 antibody
    • Memapsin-2 antibody
    • Memapsin2 antibody
    • Membrane associated aspartic protease 2 antibody
    • Membrane-associated aspartic protease 2 antibody
    • Transmembrane aspartic proteinase Asp2 antibody
    see all

画像

  • Western blot - Anti-BACE1 antibody (ab2077)
    Western blot - Anti-BACE1 antibody (ab2077)
    All lanes :

    Lane 1 : A431 cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 2 : A549 cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 3 : Caco-2 cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 4 : Daudi cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 5 : HeLa cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 6 : K562 cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 7 : Jurkat cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 8 : SK-N-SH cell lysate with Anti-BACE1 antibody (ab2077)
    Lane 9 : THP-1 cell lysate with Anti-BACE1 antibody (ab2077)

    Lysates/proteins at 15 µg per lane.

    Blocking peptides at 1 µg/ml per lane.

    Secondary
    All lanes : Rabbit IgG antibody (HRP) at 1/10000 dilution

    Developed using the ECL technique.

    Additional bands at: 65 kDa (possible glycosylated form)



    10% gel.

    Running conditions: 130v for 2 hours.

    Transfer conditions: wet, 250mA, 2 hrs (Nitrocellulose membrane).

    Blocking condition: 5% non-fat dry milk in TBS, 4C, overnight.

    Primary antibody incubation: Room temperature for 1 hour.

    Secondary antibody incubation: Room temperature for 1 hour. 

    Washing conditions: 15 mL TSBT, 3 x 10 minutes. 

    Exposure: ECL solution

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BACE1 antibody (ab2077)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BACE1 antibody (ab2077)

    Immunohistochemical analysis of paraffin-embedded mouse brain tissue using ab2077 at 2.5 ug/ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4˚C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin.

  • Immunocytochemistry/ Immunofluorescence - Anti-BACE1 antibody (ab2077)
    Immunocytochemistry/ Immunofluorescence - Anti-BACE1 antibody (ab2077)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed mouse 3T3/NIH cells labeling BACE with ab2077 at 20 ug/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI staining (blue). Image showing both membrane and cytosol staining on 3T3/NIH cells.

  • Western blot - Anti-BACE1 antibody (ab2077)
    Western blot - Anti-BACE1 antibody (ab2077)
    All lanes : Anti-BACE1 antibody (ab2077) at 1 µg/ml

    Lane 1 : Human brain tissue lysate with absence of blocking peptide
    Lane 2 : Human brain tissue
    lysate with BACE1 peptide (ab7883)
    Lane 3 : Mouse 3T3/NIH cell lysate

    Lysates/proteins at 15 µg per lane.

    Secondary
    All lanes : Goat anti-rabbit IgG HRP conjugate at 1/10000 dilution

    Observed band size: 70 kDa
    why is the actual band size different from the predicted?



    Incubate the antibody for 1 hour at room temperature in 5% NFDM/TBST.

     

     

  • Immunocytochemistry/ Immunofluorescence - Anti-BACE1 antibody (ab2077)
    Immunocytochemistry/ Immunofluorescence - Anti-BACE1 antibody (ab2077)Image courtesy of an anonymous Abreview.
    ab2077 staining BACE1 in human glioblastoma cell line D54MG by Immunocytochemistry/ Immunofluorescence.
    Cells were fixed in paraformaldehyde, permeabilized with 0.1% Triton X-100, blocked with 0.5% BSA for 20 minutes at room temperature, then incubated with ab2077 at a 1/50 dilution for 16 hours at 4°C. The secondary used was a TRITC conjugated goat anti-rabbit polyclonal, used at a 1/400 dilution. Nuclei are counterstained with DAPI.

    See Abreview

  • Immunocytochemistry/ Immunofluorescence - Anti-BACE1 antibody (ab2077)
    Immunocytochemistry/ Immunofluorescence - Anti-BACE1 antibody (ab2077)Image courtesy of an anonymous Abreview.
    ab2077 staining BACE1 in rat glioma cell line C6 by Immunocytochemistry/ Immunofluorescence.
    Cells were fixed in paraformaldehyde, permeabilized using 0.1% Triton X-100, blocked with 0.5% BSA for 20 minutes at room temperature, then incubated with ab2077 at a 1/50 dilution for 16 hours at 4°C. The secondary used was a TRITC conjugated goat anti-rabbit polyclonal, used at a 1/400 dilution. Nuclei are counterstained with DAPI.

    See Abreview

参考文献

This product has been referenced in:

  • Wu Y  et al. Microglia and amyloid precursor protein coordinate control of transient Candida cerebritis with memory deficits. Nat Commun 10:58 (2019). Read more (PubMed: 30610193) »
  • Wang W  et al. Toxic amyloid-ß oligomers induced self-replication in astrocytes triggering neuronal injury. EBioMedicine 42:174-187 (2019). Read more (PubMed: 30926423) »
See all 51 Publications for this product

ab2077 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

レビューと Q&A

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1-10 of 15 Abreviews or Q&A

Question

Thank you for your response.

Our customer has also requested for a credit note for this one too.

Appreciate your time and if you could kindly organise this for us.

Thanks again and enjoy your day J

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Answer

Thank you for contacting us.

Your credit note ID is xxxxx.

I am sorry that this antibody did not perform as stated on the datasheet, I have asked our Finance department to issue a credit note for you. The credit note may be used in one of the following ways:

(1) Redeemed against the original invoice if this hasn't already been paid.
(2) Held on the account for use against a future order.
(3) A full refund can be offered where no other invoices are outstanding.

Please contact your Finance department to confirm how you would like the credit note to be used and ensure it is not redeemed without your knowledge.

To specifically receive a refund please ask your Finance department to contact our Finance department at creditcontrol@abcam.com or by telephone using the information at the “Contact Us” link in the top right corner of our website.

The credit note ID is for your reference only, please refer to the credit note ID in any correspondence with our accounting department. We will send you the completed credit note by email or postal mail with the actual credit note number which will start with the letters CGB.

I hope this experience will not prevent you from purchasing other products from us in the future. Our Scientific Support team is always at your service should you require further expert advice

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Question

Thank you for your email.

Our customer has responded with the following;

Could you please confirm if a loading control was included and what the results were? A loading control assures that the samples used are of good quality.

My positive control were SH-SY5Y and mice brain lysate (30ug/well). As you know both are neurons and have plenty amount of BACE1.

2. We have some data that suggest that the BACE1 protein is difficult to transfer to the membrane in WB application. I would appreciate if you could confirm if the transfer of protein onto the membrane was assessed and what was the result? I have been working on BACE1 for more than 3 years, and I think this question is not true and accepteptable.

3. Could you confirm the isotype that is recognised by the secondary antibody and weather it is known to work well with other primary antibodies? It was goat Anti-Rab IgG (Total) HRP

Our all secondary antibody have been already optimized and they are working very well with 1:3000 dilution.

I look forward to your response.

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Answer

Thank you for your message and for providing this further information.

I am sorry to confirm that we can not make any further suggestions in this case to improve the results. It seems that you regrettably received a bad vial. For that reason I would be pleased to arrange a refund, credit note or free of charge replacement in compensation.

I look forward to hearing from you with details of how you would like to proceed.

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Question

Product code: 2077
Lot number: GR1723-1
Inquiry: Hi Technical, We have a customer who is experiencing problems with this antibody. Please find attached the customer's questionnaire.

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Answer

Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from our Anti-BACE1 antibody (ab2077) .

The details you have kindly provided will enable us to investigate this case for you and this is also helpful in our records for monitoring of quality.

I would like to reassure you that ab2077 is tested and covered by our 6 month guarantee for use in WB and mouse as well as human samples. In the event that a product is not functioning in the applications cited on the product data sheet, we will be pleased to provide a credit note or free of charge replacement.

Regrettably reviewing all the details there are not many suggestions we can make to improve the protocol. However, I would like to investigate the case a bit further for you. Before deciding how to proceed with this particular case, I would appreciate if you can confirm some details of the procedure.

1. Could you please confirm if a loading control was included and what the results were? A loading control assures that the samples used are of good quality.

2. We have some data that suggest that the BACE1 protein is difficult to transfer to the membrane in WB application. I would appreciate if you could confirm if the transfer of protein onto the membrane was assased and what was the result?

3. Could you confirm the isotype that is recognised by the secondary antibody and weather it is known to work well with other primary antibodies?

Thank you very much for your cooperation. Please do not hesitate to contact me again with the further requested details. I look forward to your answer and am sure we will be able to find a solution.

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Question

The ab2077 I have was giving the expected signal for my past several lysates. To verify the exact band, since I was getting one at approx 70 and another at approx 55/60, I used the mouse lysate. The strong band at 100 kD for the mouse lysate was unexpected and confusing, leading me to believe the mouse lysate wasn't good. I thought running it again with the mouse lysate in the middle of the gel might clarify where the positive control was appearing. I also thought that even if I ran this again with a new vial of ab2077, I still wouldn't have a positive control to verify my band (and appease future reviewers).

Having said that, perhaps you're right that I should at least try another ab2077. I'd rather do that then try a different primary antibody. So, go ahead and send a new ab2077. If I get the same thing, then I'll have to decide how to proceed to verify that the bands I'm seeing are indeed the glycosylated and non-glycosylated forms for BACE1.

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Answer

Thank you for your reply.

I am sending you a new vial of ab2077, your new order number is ** and you should have the new antibody by 10.30am tomorrow morning. If there is anything else I can help you with, or if you have any issues with the new vial of the antibody please let me know.

I hope your experiments go well.

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Question

I understand that you can't send all the reagents, but I've been working on this project for too long to quit. I don't think anything's wrong with the ab2077 antibody. Would you be able to send me only the 3T3 lysate? I need 30 ul to run at least 2 lanes. If that works, then I'll order (and pay for) more ab2077 to run the experiment to completion.

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Answer

Thank you for your reply.

If sending you a new aliquot of the cell lysate would help you to complete your experiments, then I would be happy to do so. However, I am confused as to why you think that the current vial of ab2077 that you have is ok when it is not giving the expected signal. I was hoping that if I send you a different antibody against the same target, then you would be able to detect your band of interest.

As I mentioned previously, I would be happy to send a new vial of cell lysate, I just wanted to confirm with you the course of action that you wish to take and to double check that you did not want to try a different primary antibody.

I look forward to your reply.

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Question

Thanks for your response. For some reason the email I sent a couple days ago bounced back. Please see attached an overlay of the ECL image with the Marker image. It definitely seems like there are two bands (between 75 kD and 50 kD) that correspond with the BACE1 protein. Specifically, one at approx 70 kD and one at approx 56 kD. Based on the data sheets you sent me, the 70 kD band is probably the glycosylated form and the smaller approx 56 band might be the unglycosylated form. It is strange that the mouse lysate appears in the 100 kD area, but there seems to be a faint band in the 3T3 lanes at approx 56 kD. I need to re-run this in the middle of a gel to be sure, but I need more lysate.

I'm not ready to jump ship and go with a different company. Is there another positive control you'd recommend? Also, since the peptide hasn't worked, I'm guessing the batch was somehow bad. I also think I should try a new BACE1 antibody along with the ab2077 I still have. So, can you please send me the following: 1) peptide ab7883, 2) either a different positive control or more of the mouse lysate 3T3 (at least 40ul), 3) ab108394 (since perhaps a monoclonal Ab would decrease the non-specific bands I'm seeing).

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Answer

Thank you for your reply.

I am sorry about the issues you have been having with ab2077 in western blot. I cannot send you all of the reagents that you are requesting, I can either send 1 free antibody as a replacement for ab2077, or I can process a refund for the cost of the original purchase.

Please let me know how you would like to proceed.

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Question

Not seeing the correct size band in wetern blotiing

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Answer

Thank you for calling Abcam yesterday.

I am sorry about the issues you have been having with ab2077 in western blot. Having talked to my colleagues, I think the best way to move forward with this is to either replace the antibody that you currently have or to refund the cost of the original purchase.

I do not think that there are any protocol tips that I could give that would help resolve the banding pattern that you are seeing and can only assume that the vial of ab2077 that you received is not working correctly.

A few alternative BACE1 antibodies to consider are:

https://www.abcam.com/BACE1-antibody-EPR3956-ab108394.html
https://www.abcam.com/BACE1-antibody-ab63954.html



If either of these antibodies would work for you (or if there is another BACE1 antibody that you would like to try), please let me know and I will be happy to send it out to you.

I look forward to your reply.

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Question

Called because product was giving non specific bands

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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 the free of charge blocking peptide for this product with the order number I have also included our protocol for blocking with the immunizing peptide for your review. 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.

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Immunocytochemistry/ Immunofluorescence abreview for Anti-BACE1 antibody

 Good
Abreviews
abreview image
Application
Immunocytochemistry/ Immunofluorescence
Sample
Rat Cell (murine glioma cell line C6)
Specification
murine glioma cell line C6
Fixative
Paraformaldehyde
Permeabilization
Yes - 0,1% Triton X 100
Blocking step
BSA as blocking agent for 20 minute(s) · Concentration: 0.5% · Temperature: RT°C
Read More

Abcam user community

Verified customer

投稿 Aug 30 2011

Immunocytochemistry/ Immunofluorescence abreview for Anti-BACE1 antibody

 Average
Abreviews
abreview image
Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (human glioblastoma cell line D54MG)
Specification
human glioblastoma cell line D54MG
Fixative
Paraformaldehyde
Permeabilization
Yes - 0,1% Triton X 100
Blocking step
BSA as blocking agent for 20 minute(s) · Concentration: 0.5% · Temperature: RT°C
Read More

Abcam user community

Verified customer

投稿 Aug 30 2011

1-10 of 15 Abreviews or Q&A

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