重组
RabMAb

Anti-Polyethylene glycol抗体[PEG-B-47] (ab51257)

概述

  • 产品名称
    Anti-Polyethylene glycol抗体[PEG-B-47]
    参阅全部 Polyethylene glycol 一抗
  • 描述
    兔单克隆抗体[PEG-B-47] to Polyethylene glycol
  • 宿主
    Rabbit
  • 特异性
    This antibody recognizes the terminal methoxy group of the PEG molecule. This antibody does not cross react with non-specific targets in blood or serum. Both free form of PEG (unconjugated version) and conjugated PEG were tested by sandwich ELISA using ab51257; results showed that ab51257 only detects conjugated forms of PEG. To detect free form of PEG, we recommend using ab133471. ab133471 recognizes PEG backbone and that PEG is not required to be bound to other molecules.
  • 经测试应用
    适用于: ELISA, WB, IHC-Pmore details
  • 免疫原

    Chemical/ Small Molecule corresponding to Polyethylene glycol conjugated to Keyhole Limpet Haemocyanin (KLH). KLH-Polyethylene glycol with a terminal methoxy group.

  • 常规说明

    Abcam recommended secondaries - Goat Anti-Rabbit HRP (ab205718) and Goat Anti-Rabbit Alexa Fluor® 488 (ab150077).

    See other anti-rabbit secondary antibodies that can be used with this antibody.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    This product is a recombinant rabbit monoclonal antibody.

性能

  • 形式
    Liquid
  • 存放说明
    Shipped at 4°C. Upon delivery aliquot. Store at -20°C.
  • 存储溶液
    pH: 7.40
    Preservative: 0.01% Sodium azide
    Constituents: 40% Glycerol, 0.05% BSA, 59% PBS
  • Concentration information loading...
  • 纯度
    Protein A purified
  • 克隆
    单克隆
  • 克隆编号
    PEG-B-47
  • 同种型
    IgG
  • 研究领域

应用

Our Abpromise guarantee covers the use of ab51257 in the following tested applications.

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

应用 Ab评论 说明
ELISA Use a concentration of 0.5 - 5 µg/ml.
WB Use a concentration of 0.015 - 0.075 µg/ml.
IHC-P 1/100.

This antibody only works on the tissues when the animals are injected with a PEGylated protein.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

靶标

  • 别名
    • PEG antibody

图片

  • Immunohistochemistry images of anti-PEG (brown) and hematoxylin counterstaining (blue) of tumor (A1–A3 and B1–B3) and muscle tissue (A4 and B4) 30 minutes (A) and 2 h (B) post injection of Spago Pix

    Tumor tissue at both time points is shown at three different enlargements where the enlarged areas are indicated by the square boxes. Vessels (V), fibroblasts (Fb) and fat cells (Fc) are indicated with rounded lines. Note that the tumor tissue in panel B1–B3 originates from a mouse injected with a dose of 10 µmol Mn/kg, whereas the other tissues originate from animals administered 20 µmol Mn/kg.

    Paraformaldehyde fixed muscle and tumor tissues collected before injection and at 30 minutes and 2–4 h post-dose were paraffin embedded and thereafter cut (4 µm) and positioned on glass tissue slides. Prior to staining, the slides were placed in an antigen-retrieval solution using an automated pre-treatment module (PT-Link; Dako, Glostrup, Denmark). Slides were stained for PEG in an automated immunohistochemistry robot (Autostainer; Dako). PEG is detected with ab51257 at 0.6 μg/ml.

    (From Figure 6 Eriksson et al)

  • Anti-Polyethylene glycol antibody [PEG-B-47] (ab51257) at 0.015 µg/ml + BSA-PEG at 10 µg

    Secondary
    Goat Anti-Rabbit IgG, (H+L), HRP-conjugated at 1/1000 dilution


    Blocking buffer and concentration: 5% NFDM/TBST.

    Diluting buffer and concentration: 5% NFDM /TBST.

  • Comparison of sandwich ELISA using RabMAb/RabMAb (ab51257/ab51257) and MAb/RabMAb (Vendor A/ab51257) for capture/detection.

    Ab51257/ab51257*: Plate coated with 5 ug/mL of #47; 5 ug/mL of #47 used for detection (*Anti-PEG 47 biotin labeled)

    Vendor A/ ab51257*: Plates coated with 100 ug/mL of Vendor A Mouse MAb; 5 ug/mL of #47 used for detection (*Anti-PEG 47 biotin labeled)

  • Immunoblot of different antigen-Polyethlene glycol preparations.

    Western Blot images using ab51257, unpurified, to detect BSA-PEG.

    Lane 1: 300 ng/ml;
    Lane 2: 75 ng/ml;
    Lane 3: 15 ng/ml;
    Lane 4: 1.5 ng/ml.

  • Comparison of 10 ug/well of activated linear (PEG5K) and branched (PEG40K) PEG using 5 ug/ml of ab51257 in Direct ELISA assay.

    Accuracy: By detecting the methoxy group of the PEG molecule itself, ab51257 is useful in measuring the pharmacokinetics of PEG-modified molecules in vivo. Data indicate that ab51257 detects various length Y-chain PEG molecules as well as single chain PEG molecules with equal affinity. Ab51257 does not cross react with non-specific targets in blood or serum.

  • ab51257, unpurified,  staining mouse kidney at 1:10 dilution. Left panel: without animal injection with a PEGylated protein. Right panel: with animal injection with a PEGylated protein.

  • Comparison of ab51257 and Vendor A mouse MAb in Direct ELISA assy.

    Goat anti-rabbit IgG-AP used for anti-PEG-47 detection; goat anti-mouse IgM-AP used for Vendor A MAb detection.

    Fig 1a. Direct ELISA using 1 ug/mL of PEG-GCSF.

    Fig 1b. Direct ELISA using 1 ug/mL of PEG-IFN.

    Comparison to other anti-PEG: In both direct and sandwich ELISA assays, ab51257 shows greater affinity and accuracy than other anti-PEG antibodies when determining the concentration of PEG or PEG-modified proteins. Results were similar whether detecting PEG itself or PEG-modified targets.

  • ELISA assay using ab51257 to detect different forms of PEG. PEG40000-BSA is a 40 kDa PEG molecule attached to BSA. PEG5000-BSA is a 5 kDa linear PEG molecule attached to BSA.
  • ab51257, unpurified, staining mouse muscle at 1:10 dilution. Left panel: without animal injection with a PEGylated protein. Right panel: with animal injection with a PEGylated protein.

  • ab51257, unpurified,  staining mouse liver at 1:10 dilution. Left panel: without animal injection with a PEGylated protein. Right panel: with animal injection with a PEGylated protein.

  • ab51257, unpurified, staining mouse spleen at 1:10 dilution. Left panel: without animal injection with a PEGylated protein. Right panel: with animal injection with a PEGylated protein.

  • Sandwich ELISA-Left Graph generated using unpurified ab51257 and right using purified ab51257 at 1μg/mL. Antigen concentration range 0.0156 - 1 μg/mL for PEG-IFN-α-2b. Secondary antibody was an Alkaline Phosphatase-conjugated Goat Anti-Rabbit IgG(H+L) at 1/2500.

  • Direct ELISA-Left Graph generated using unpurified ab51257 and right using purified ab51257 at 1μg/mL. Antigen concentration range 0.0039 - 0.25μg/mL for PEG-BSA. Secondary antibody was an Alkaline Phosphatase-conjugated Goat Anti-Rabbit IgG(H+L) at 1/2500.

  • Direct ELISA-Left Graph generated using unpurified ab51257 and right using purified ab51257 at 1μg/mL. Antigen concentration range 0.0156 - 1 μg/mL for ADI-PEG. Secondary antibody was an Alkaline Phosphatase-conjugated Goat Anti-Rabbit IgG(H+L) at 1/2500.

  • Direct ELISA-Left Graph generated using unpurified ab51257 and right using purified ab51257 at 1μg/mL. Antigen concentration range 0.0078 - 0.5 μg/mL for both PEG5000 and PEG20000. Secondary antibody was an Alkaline Phosphatase-conjugated Goat Anti-Rabbit IgG(H+L) at 1/2500.

文献

This product has been referenced in:
  • Chen YF  et al. Disulfide-cross-linked PEG-block-polypeptide nanoparticles with high drug loading content as glutathione-triggered anticancer drug nanocarriers. Colloids Surf B Biointerfaces 165:172-181 (2018). IHC . Read more (PubMed: 29482128) »
  • Triplett TA  et al. Reversal of indoleamine 2,3-dioxygenase-mediated cancer immune suppression by systemic kynurenine depletion with a therapeutic enzyme. Nat Biotechnol 36:758-764 (2018). Read more (PubMed: 30010674) »
See all 39 Publications for this product

客户评价及客户问答

1-10 of 10 Abreviews or Q&A

Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Macrophages)
Permeabilization
Yes - 0.3% Triton X-100
Specification
Macrophages
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 21°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

提交于 Nov 08 2016

Answer

In our validation of ab51257 in IHC-P, antigen retrieval was not necessary. ab51257 was validated internally with mice injected intraperitoneally using 100 ug of BSA-PEG as an antigen. Heart, kidney, liver and spleen tissue were stained 4 hours post-injection using Anti-PEG RabMAb without antigen retrieval.



However, some of the papers that have used the antibody in IHC-P did use an antigen retrieval step (one example used a head mediated retrieval on a DAKO autostainer). A list of publications that used the antibody in IHC are listed below, which may be of further assistance:



http://www.ncbi.nlm.nih.gov/pubmed/24749870?dopt=Abstract

http://www.ncbi.nlm.nih.gov/pubmed/25296030?dopt=Abstract

http://www.ncbi.nlm.nih.gov/pubmed/23665258?dopt=Abstract

http://www.ncbi.nlm.nih.gov/pubmed/23788571?dopt=Abstract

http://www.ncbi.nlm.nih.gov/pubmed/22745537?dopt=Abstract

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Question
Answer

The antibody ab51257 is specific to the terminal of PEG molecule and is made up of rabbit IgG. In contrast, ab133471 is specific to the backbone of the PEG molecule and is made up of rabbit monoclonal IgM.

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Question
Answer

Both antibodies ab51257 and ab53449 are the same clone (PEG-B-47). Product ab53449 is the biotinlyated version of ab51257.

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Question
Answer

Yes, ab51257 is used in the kit as the capture antibody in the kit and it has been shown to detect both linear and branched PEG.

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Question
Answer

Both ab51257 and ab53449 detect linear and branched PEG (based on our in-house ELISA results).

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Answer


Both free form of PEG (unconjugated version) and conjugated PEG were tested by sandwich ELISA using this anti-PEG antibody (ab51257); however, our results showed that this antibody does not detect unconjugated PEG.

If you need an antibody which detects the free form of PEG, I would recommend using an anti-PEG Backbone antibody, ab133471. This antibody recognizes PEG backbone and that PEG is not required to be bound to other molecules.

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Question
Answer

We used PEG with MW of 5K to generate ab51257. The smallest PEG molecule which our antibody can detect is PEG750. Based on customer feedback, the biggest PEG molecule detected is PEG 40K. Additionally, this product has been shown to detect both linear and branched PEG.

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Answer

Thank you for contacting us and your interest in our products.

The Anti-Polyethylene glycol antibody [PEG-B-47] (catalogue number ab51257, 2061-1) has been used in IHC and is expected to be able to detect PEG if present in the tissue with IHC of formalin fixed, paraffin embedded sections.

This was performed at Epitomics with mice injected intraperitoneally using 100 ug of BSA-PEG as an antigen. Heart, kidney, liver and spleen tissue were stained 4 hours post-injection using Anti-PEG RabMAb (Cat #2061-1). Attached please find images of the results obtained.

I hope this information has been helpful. If I can be of any further assistance, please do not hesitate to let me know.

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Question
Answer

Thank you for your enquiry regarding ab51257.

I have spoken to my colleagues in the Lab and they have confirmed me that the concentration is batch dependent but the actual concentration of this product is printed on the label of the vial.

If you need any further assistance, please do not hesitate to contact me.

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

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