Anti-BSD antibody (ab38307)

Reviews (1)Q&A (4)References (2)

Overview

  • Product name

    Anti-BSD antibody

  • Description

    Rabbit polyclonal to BSD

  • Host species

    Rabbit

  • Specificity

    The antibody detects a 13 kDa protein corresponding to the molecular mass of Bsd on SDS-PAGE immunoblots of 293 cells transfected with a lentiviral vector that expresses the Bsd gene product, but is not detected in mock transfected cells.

  • Tested applications

    Suitable for: WB, ELISA, ICC/IFmore details

  • Species reactivity

    Reacts with: Species independent

  • Immunogen

    Synthetic peptide conjugated to KLH corresponding to amino acid residues in the N-terminal region of Blasticidin S Deaminase from Aspergillus Terrus.

Properties

  • Form

    Liquid

  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.

  • Storage buffer

    Preservative: 0.05% Sodium azide
    Constituents: 0.1% BSA, 50% Glycerol, PBS
  • Concentration information loading...

  • Purity

    Immunogen affinity purified

  • Purification notes

    This antibody was affinity purified using the N-terminal Bsd peptide (without carrier).

  • Clonality

    Polyclonal

  • Isotype

    IgG

  • Research areas

Applications

Our Abpromise guarantee covers the use of ab38307 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
WB 1/2000. Detects a band of approximately 13 kDa.
ELISA 1/4000.
ICC/IF 1/100.

Target

  • Relevance

    Retroviral vectors are commonly used for gene expression in vitro and in vivo. The structure of these vectors usually consists of one gene of interest and a selectable marker gene. Fast selection without damaging cells is a critical step for significant gene expression. Blasticidin S deaminase (Bsd) isolated from Bacillus cereus has a neutralizing action on the highly toxic antibiotic blasticidin. A variety of commercially available retroviral vectors, such lentiviral expression systems, utilize Bsd expression for selection using blasticidin. This selectable marker allows selection of stable mammalian cell lines in as little as one week.

  • Alternative names

    • Blasticidin-S deaminase antibody
    • BSD antibody

Images

  • Western blot - Anti-BSD antibody (ab38307)
    Western blot - Anti-BSD antibody (ab38307)
    All lanes : Anti-BSD antibody (ab38307)

    Lane 1 : 293 cells mock transfected
    Lanes 2-3 : 293 cells transiently transfected with pLenti6/TR lentiviral vector

    Observed band size: 13 kDa
    why is the actual band size different from the predicted?



    Each lane was loaded with 10-20µg total protein.
  • Immunocytochemistry/ Immunofluorescence - Anti-BSD antibody (ab38307)
    Immunocytochemistry/ Immunofluorescence - Anti-BSD antibody (ab38307)Image courtesy of Charles Mashburn and Dr. George Smith at the University of Kentucky, Spinal Cord and Brain Injury Research Center
    ab38307, staining BSD in 293 cells mock transfected (left) and transiently transfected with pLenti6/TR lentiviral vector (right). The cells were fixed in paraformaldehyde (3.75%) for 30 min., then permeabilized in 0.2% Triton X-100 for 15 min. Blocking was done using 1% BSA for 30 min. The primary was used at 1/100 and then an appropriate secondary antibody conjugated to Texas Red was used.

References

This product has been referenced in:

  • Jammart B  et al. Very-low-density lipoprotein (VLDL)-producing and hepatitis C virus-replicating HepG2 cells secrete no more lipoviroparticles than VLDL-deficient Huh7.5 cells. J Virol 87:5065-80 (2013). Read more (PubMed: 23427158) »
  • de Wispelaere M & Yang PL Mutagenesis of the DI/DIII linker in dengue virus envelope protein impairs viral particle assembly. J Virol 86:7072-83 (2012). Read more (PubMed: 22532681) »
See all 2 Publications for this product

Publishing research using ab38307? Please let us know so that we can cite the reference in this datasheet.

Customer reviews and Q&As

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

 Poor
Abreviews
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Fibroblasts transduced with BSD-resistance gene)
Specification
Fibroblasts transduced with BSD-resistance gene
Fixative
Paraformaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: 0.01M citrate (pH 6.)
Permeabilization
No
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
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Abcam user community

Verified customer

Submitted Oct 08 2012

Question

Would like to test this antibody in IHC-P.

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Answer

DISCOUNT CODE: ***
Expiration date: October 5, 2012

This code will give you 1 free primary antibody before the expiration date. To redeem this offer, please submit an Abreview forab38307 in IHC-Pand 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.

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.

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Question

Would like to try this antibody in IHC on paraffin-embedded tissue.

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Answer

Thanks for your call today and for your interest in ab38307. Please see below for your code to test this antibody in paraffin embedded tissue staining (IHC-P) and let me know if you have any questions.

DISCOUNT CODE: ***
Expiration date: June 25, 2012

This code will give you 1 free primary antibody before the expiration date. To redeem this offer, please submit an Abreview for ab38307 in IHC-P and 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.

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.

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Question

Please see below the details of customer's complaint. Attached is the image of the gel. Antibody code: ab38307 Batch number: GR23921 Antibody storage conditions (temperature/reconstitution etc): -200C Description of the problem (high background, wrong band size, more bands, no band etc.): no bands Sample (Species/Cell extract/Nuclear extract/Purified protein/Recombinant protein etc.): Plasmodium falciparum - Cell extract. Sample preparation (Buffer/Protease inhibitors/Heating sample etc.): A fresh protein lysate was used each time, all procedure was performed on ice/40C, Parasites Cells were diluted in 96ul of ice cold PBSx1 and 4ul of protease inhibitor x25, after a 10min incubation 100ul of a 700C pre warmed sample buffer was added. Sample was place in 700C and after a few trails in 950C for 5min before loaded..   Amount of protein loaded: 10-20ul (concentration was not measured because the cell extract is not purified) Electrophoresis/Gel conditions (Reducing or Non-reducing gel, % of the gel etc.) Bio-Rad 4-20% gel  (Mini-PROTEAN) Transfer and blocking conditions (Buffer/time period, Blocking agent etc.) Transfer: transfer buffer - 700ml DDW, 200ml methanol and 100ml transfer bufferx10 (Tris 478.8mM, Glycin 386.3mM, SDS 0.37%, Ph 8.3). 130 mA for 90min or 60min. Blocking: different blocking buffers were used: 5% skim milk or 5% BSA in 100ml PBS-T (0.1% tween or 0.04% tween) for 1h at room temp or overnight at 40C. Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step) Rabbit polyclonal to BSD (abcam ab38307), different ab dilutions were used: 1:2000 or 1:1000 in 5% skim milk 0.1% tween or in 1% skim milk and 0.04% tween. Incubation times: on shaker- 1h at room temp or over night at 40C. Different washes were preformed in PBS-T (0.1% tween): on shaker, 3 times x 10min or 3 times x 3min.  Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step) Goat anti-rabbit IgG (H+L). different ab dilutions were used: 1:10000 in 5% skim milk 0.1% tween or in 1% skim milk and 0.04% tween. Incubation times: on shaker- 1h at room. Different washes were preformed in PBS-T (0.1% tween): on shaker, 3 times x 10min or 3 times x 3min.  Detection method (ECL, ECLPlus etc.): Easy-ECL (Pierce). Exposure time at least 20min at room temp or overnight in 40C Positive and negative controls used (please specify): Positive control – a transgenic parasites line that is expressing BSD and is grown on 2ug/ml blasticidin. Negative control - a blasticidin non resistant parasite line. (Cannot grow on 2ug/ml blasticidin). Optimization attempts (problem solving) How many times have you tried the Western? 10 Have you run a "No Primary" control? No Do you obtain the same results every time? Yes, no bend except the ladder. What steps have you altered? Different loading volume of  highly prodused sample, transfer time, blocking buffer and time, primary antibody dilutions and incubation times, wash times, secondary antibody dilutions and incubation times and different exposure times  (as specified above).

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Answer

Thank you for contacting us. I am sorry to hear you have been experiencing problems with ab38307. The customer has made many changes to optimize and it is disappointing that the antibody is still not working. I would be happy to refund or replace the product. The only recommendation I would like to make is using 10% saponin for lysis, reviewing online protocols this is a common method to lyse Plasmodium falciparum and may be more effective than 1xPBS only. I hope this information helps. Looking forward to your reply on how to proceed.

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Question

Could you tell me if the BSD antibody (ab38307) works on cells fixed with paraformaldehyde? Thank you very much in advance,

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Answer

Thank you for your inquiry. The cells in the ICC/IF image for ab38307 were fixed in paraformaldehyde (3.75%) for 30 min., then permeabilized in 0.2% Triton X-100 for 15 min. Blocking was done using 1% BSA for 30 min, then the labeling was performed as described in the caption. I have updated our image caption with this information. Please contact us with any other questions.  

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