Native Clostridium difficile Toxin B protein (ab124001)

Reviews (1)Q&A (5)References (2)

Overview

  • Product name

    Native Clostridium difficile Toxin B protein

  • Protein length

    Full length protein

Description

  • Nature

    Native

  • Source

    Native
  • Amino Acid Sequence
    • Accession
      P18177

    • Molecular weight

      270 kDa

    • Amino acids

      2 to 2366

Associated products

Specifications

Our Abpromise guarantee covers the use of ab124001 in the following tested applications.

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

  • Applications

    SDS-PAGE

    Functional Studies

  • Purity

    > 95 % SDS-PAGE.
    Clostridium difficile Toxin B has been highly purified.

  • Form

    Lyophilised

  • Additional notes

    This protein is derived from a pathogenic organism, and may be involved in a disease process, consequently exposure may have adverse health effects.

    Reconstituted liquid can be stored at -80°C or -20°C to extend stability.

    Functional Studies Protocol

    1.1 Vero cells are plated out the night before on a 96-well plate at a concentration of 20K cells per well (in 100ml), then incubated at 37°C, 5% CO2. The following day they should be lightly confluent.
    1.2 Lyophilised toxin/toxoid is resuspended as per the instructions in the certificate of analysis, and diluted in duplicate to 0.1mg/ml in DMEM (200ml total) without FBS in the first well of a non-TC-treated 96-well plate. A serial 5-fold dilution is performed (40ul in 160ul) in the same media using a multichannel for 11 dilutions in total, leaving the 12th dilution as a media control.
    1.3 20ul of each dilution is then pipetted onto each well of the pre-prepared Vero cells and then incubated at 37°C, 5% CO2 for 24 hours.
    1.4 The following day, the cells are examined using an inverted phase contrast tissue culture microscope. Cells with active toxin on should be rounded, cells without significant toxicity should look like the control cells.

  • Concentration information loading...

Preparation and Storage

  • Stability and Storage

    Shipped at 4°C. Store at +4°C. Avoid freeze / thaw cycle. Please see notes section.

    Constituents: 1.19% HEPES, 5% Sucrose, 0.88% Sodium chloride

  • Reconstitution
    Reconstitute in 250µl sterile distilled water. Reconstituted liquid can be stored at -80°C or -20°C to extend stability.

General Info

  • Alternative names

    • C. difficile Toxin B
    • Clostridium difficile Toxin B
    • Cytotoxin B
    • tcdB
    • toxB
    • Toxin B
    see all

  • Relevance

    Clostridium difficile Toxin B (tcdB or ToxB) is a glucosyltranferase which is known to innactivate Rho, Cdc42 and Rac within target cells. This toxin is encoded on a pathogenicity region of the C. difficile chromossome and is expressed during the log and stationary phases of growth in response to a variety of environmental stimuli.

  • Cellular localization

    Secreted

Images

  • SDS-PAGE - Native Clostridium difficile Toxin B protein (ab124001)
    SDS-PAGE - Native Clostridium difficile Toxin B protein (ab124001)
    SDS-PAGE showing ab124001 at 270 kDa.

References

This product has been referenced in:

  • Hartley-Tassell LE  et al. Lectin activity of the TcdA and TcdB toxins of Clostridium difficile. Infect Immun N/A:N/A (2018). Read more (PubMed: 30530621) »
  • Hutton ML  et al. Bovine antibodies targeting primary and recurrent Clostridium difficile disease are a potent antibiotic alternative. Sci Rep 7:3665 (2017). Neutralising ; Clostridium difficile . Read more (PubMed: 28623367) »
See all 2 Publications for this product

Publishing research using ab124001? Please let us know so that we can cite the reference in this datasheet.

Customer reviews and Q&As

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1-6 of 6 Abreviews or Q&A

Inositol hexakisphosphate (IP6)-induced autoprocessing

 Excellent
Abreviews
abreview image
Application
Functional Studies
The TcdB undergoes auto-processing in the presence of 1mM of IP6 as see by gel electrophoresis (see image attached).

Abcam user community

Verified customer

Submitted Jul 19 2019

Question

What C. difficile strain was used to generate purified Toxin A (cat#ab123999) and Toxin B (cat#ab124001)?”

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Answer



These products were purified from Clostridium difficile strain VPI10463 (which produces toxinotype 0 toxins). The toxins are active, each batch is tested by verocytotoxicity assay and demonstrated to be toxic to cells down to at least 5 ng/ml for toxin A and 0.05 ng/ml for toxin B. No other biological tests are performed on these toxins.

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Question

Hello,
I am potentially interested in buying some C.diff toxin but have a few
questions:
1) How long does the Toxin remain stable once reconstituted?
2) Can tha Toxin be frozen (either at -20 or -80) once reconstituted
for longterm storage?

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Answer

Thank you for contacting us.

In response to your questions:

1)The toxin is guaranteed to be stable for at least 1 month after reconstitution, (we have data that shows 6 weeks is still fine).

2)The toxin can be frozen for long term storage, but every freeze thaw will result in more and more precipitation and loss of function. I would recommend aliquotting and freezing at -80C immediately after reconstitution, then only using each aliquot once.

I hope this helps. Please do not hesitate to contact me back for further assistance.

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Question

Inquiry: Dear Sirs, Can this product, Cl.diff. Toxin B Protein ab12401, also be used as a positive control in a Cytotoxicity Assay (measuring cytopathic effect in cell culture)? Thank you and kind regards

Read More
Answer

Thank you for contacting us again.

I am delighted to send you the protocol of the cell toxicity assay with Vero cells





1 Verocycotoxicity Assay
1.1 Vero cells are plated out the night before on a 96-well plate at a concentration of 20K cells per well (in 100ml), then incubated at 37°C, 5% CO2. The following day they should be lightly confluent.
1.2 Lyophilised toxin/toxoid is resuspended as per the instructions in the certificate of analysis, and diluted in duplicate to 0.1mg/ml in DMEM (200ml total) without FBS in the first well of a non-TC-treated 96-well plate. A serial 5-fold dilution is performed (40ul in 160ul) in the same media using a multichannel for 11 dilutions in total, leaving the 12th dilution as a media control.
1.3 20ul of each dilution is then pipetted onto each well of the pre-prepared Vero cells and then incubated at 37°C, 5% CO2 for 24 hours.
1.4 The following day, the cells are examined using an inverted phase contrast tissue culture microscope. Cells with active toxin on should be rounded, cells without significant toxicity should look like the control cells.
Of course the protocol dilutions may be altered as desired by the customer to suit their specific application.

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Question

Can this product, Cl.diff. Toxin B Protein ab12401, also be used as a positive control in a Cytotoxicity Assay (measuring cytopathic effect in cell culture)? Thank you and kind regards

Read More
Answer

I have contacted the lab regarding your question and here is their answer:

Yes, it is fully cytotoxic and has been quality controlled in a vero cell cytotoxicity assay.

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Question

1) Proteine (Toxine A+B): Welcher C.difficile-Stamm? Aktivitaet?
2) Antikoerper dagegen, die NICHT am C-Terminus binden?

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Answer

Vielen Dank für Ihren Anruf.

Zu den Toxinen kann ich Ihnen folgende Informationen aus dem Labor weiterleiten:

Clostridium difficile Toxin A Protein ab123999und Clostridium difficile Toxin B Protein ab124001werden aus demClostridium difficile Stamm VPI10463 aufgereinigt, der Toxinotyp 0-Toxine produziert.

Beide Toxine sind aktiv; jeder Batch wird im Verocytotoxicity Assay getestet und ist toxisch für Zellen bis zu einer Konzentration von mindestens5 ng/ml für Toxin A und 0.05 ng/ml für Toxin B. Andere biologische Assays werden nicht durchgeführt.

Bezüglich der Antikörper, die wir gegen das Toxin A bzw. B verfügbar haben, habe vermutlich weniger gute Nachrichten für Sie:

Wir haben derzeit vier anti-Toxin A-Antikörper in unserem Katalog:

ab35021 und ab93318: Kaninchen-polyklonale, deren Immunogen allerdings C-terminal lokalisiert ist.
https://www.abcam.com/index.html?datasheet=35021 (or use the following: https://www.abcam.com/index.html?datasheet=35021).
https://www.abcam.com/index.html?datasheet=93318 (or use the following: https://www.abcam.com/index.html?datasheet=93318).

ab19953 und ab82285: Maus-monoklonale, die gegen das Volllängenprotein generiert wurden. Das Epitop wurde leider nicht kartiert. Es ist also nicht auszuschließen, dass diese Antikörper den C-Terminus binden und blockieren.
https://www.abcam.com/index.html?datasheet=19953 (or use the following: https://www.abcam.com/index.html?datasheet=19953).
https://www.abcam.com/index.html?datasheet=82285 (or use the following: https://www.abcam.com/index.html?datasheet=82285).

Wir haben derzeit drei anti-Toxin B-Antikörper in unserem Katalog:

ab77582 und ab77583: Maus-monoklonale (wie oben), die gegen das Volllängenprotein generiert wurden. Das Epitop wurde leider nicht kartiert. Es ist also nicht auszuschließen, dass diese Antikörper den C-Terminus binden und blockieren.
https://www.abcam.com/index.html?datasheet=77582 (or use the following: https://www.abcam.com/index.html?datasheet=77582).
https://www.abcam.com/index.html?datasheet=77583 (or use the following: https://www.abcam.com/index.html?datasheet=77583).

ab83066: Kaninchen-polyklonaler, der gegen das N-terminale 60 kDa katalytische Fragment generiert wurde.
https://www.abcam.com/index.html?datasheet=83066 (or use the following: https://www.abcam.com/index.html?datasheet=83066).

Ist da vielleicht einer (ab83066?) dabei, der für Ihre Versuchsvorhaben in Frage käme?

Nähere Informationen sowie die getesteten Anwendungen und Spezies können Sie auf unseren Produktdatenblättern einsehen. Wir garantieren, dass das Produkt so funktioniert wieauf dem Datenblatt steht (https://www.abcam.com/Abpromise).

Ich hoffe, dies hilft Ihnen weiter. Bitte zögern Sie nicht, sich wieder bei uns zu melden, falls Sie weitere Fragen haben.

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