Leishmania LPG Antibody | CA7AE

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Leishmania LPG Antibody | CA7AE gallery image 1

Published investigator image:
Recruitment of Syt V is impaired on phagosomes containing LPG-coated Zymosan. A and B, BMM were allowed to internalize Zym or LPG-Zym during 15 min (A) or 2 h (B), and prepared for confocal analysis. Presence (left y axis) and P/C ratio levels (right y axis) were determined for EEA1 (A) or LAMP-1 (B). C and D, BMM cells were allowed to internalize Zym or LPG-Zym for 10 min, 30 min or 2 h, fixed and stained for either endogenous Syt V (green) and LPG (red). The presence of Syt V and LPG on phagosomes is illustrated by confocal images (C). D. Quantification of Syt V recruitment (left panel) and relative Syt V levels (P/C ratio) on these phagosomes (right panel) were determined. E. Syt V-GFP cells were allowed to internalize Zym or LPG-Zym for 10 min, 30 min or 2 h, fixed and stained for LPG. Quantification of Syt V- recruitment (left panel) and relative Syt V levels on these phagosomes (right panel) were determined. The recruitment of EEA1, LAMP1 and Syt V was determined on at least 100 phagosomes for each condition, at least three independent experiments were performed and the bars show the standard deviations of one representative triplicate (*, p=0.05; **, p=0.005). Bar, 3 µm.

From: Vinet AF, Fukuda M, Turco SJ, Descoteaux A (2009) The Leishmania donovani Lipophosphoglycan Excludes the Vesicular Proton-ATPase from Phagosomes by Impairing the Recruitment of Synaptotagmin V. PLoS Pathog 5(10): e1000628.

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Leishmania LPG Antibody | CA7AE gallery image 2

Published investigator image:
Exclusion of Syt V is restricted to the LPG insertion sites on the phagosome membrane. A and B, BMM were allowed to internalize Zym or Zym-LPG during 30 min or 2 h, and stained for Syt V (green) and LPG (red). Green arrowheads indicate a localized Syt V recruitment on phagosome membrane and red arrowheads indicate a localized LPG insertion into phagosome membrane (A). C and D, Syt V-GFP cells were allowed to internalize Zym or Zym-LPG for 30 min or 2 h, fixed and stained for LPG (red). Green arrowheads indicate a localized Syt V-GFP recruitment on phagosome membrane and red arrowheads indicate a localized LPG insertion into phagosome membrane (C). A rim around a representative phagosome formed in BMM (B) or in SytV-GFP cells (D) from A and C respectively, was manually traced with a one pixel width and fluorescence intensity profile of Syt V in green and LPG in red were represented in a graph for each phagocytosis time point. Bar, 3 µm.

From: Vinet AF, Fukuda M, Turco SJ, Descoteaux A (2009) The Leishmania donovani Lipophosphoglycan Excludes the Vesicular Proton-ATPase from Phagosomes by Impairing the Recruitment of Synaptotagmin V. PLoS Pathog 5(10): e1000628.

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Leishmania LPG Antibody | CA7AE gallery image 3

Published investigator image:
Recruitment of Syt V to GM1-containing microdomains of phagosome membranes is prevented by LPG. A, BMM were allowed to internalize Zym for 30 min, fixed and stained for endogenous Syt V (green) and GM1 (red). White arrowheads indicate examples of colocalization between Syt V and GM1-positive microdomains, indicating a Syt V enrichment on these microdomains. B and C, BMM were either left untreated or treated with 10 mmol/L MßCD for 1 h before the internalization of Zym for 30 and 120 min. Cells were then fixed and stained for Syt V and LAMP-1. Representative confocal images of Syt V recruitment on cells with or without MßCD treatment is presented (B), white arrowheads indicate phagosomes. Syt V acquisition is expressed as a percentage of phagosome recruitment for Syt V. At least 100 phagosomes for each condition were assessed. Three independent experiments were performed and the bars show the standard deviations of one representative triplicate (C) (**, p=0.005). D, BMM were allowed to internalize Zym-LPG for 30 min, fixed and stained for LPG (green) and GM1 (red). White arrowheads indicate a colocalization between LPG and GM1-positive rafts. BMM were allowed to internalize Zym (E, upper panel) or LPG-Zym (E, lower panel) for 30 min, fixed and stained for Syt V (blue), LPG (green) and GM1 (red). Blue arrowheads indicate a local Syt V acquisition on phagosome membrane and yellow arrowheads indicate a local colocalization between GM1 microdomains and LPG. A rim around each phagosome was manually traced with a one pixel width and fluorescence intensity profile of Syt V in blue, LPG in green and GM1 in red were represented in a graph (F). Bars, 3 µm (A, B and D) or 1 µm (E).

From: Vinet AF, Fukuda M, Turco SJ, Descoteaux A (2009) The Leishmania donovani Lipophosphoglycan Excludes the Vesicular Proton-ATPase from Phagosomes by Impairing the Recruitment of Synaptotagmin V. PLoS Pathog 5(10): e1000628.

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Leishmania LPG Antibody | CA7AE gallery image 4

Published investigator image:
The inhibition of the V-ATPase acquisition on phagosomes is specific for the promastigote stage. A–C, BMM cells were infected with either WT promastigotes or amastigotes for 2 h and 24 h, fixed and stained for V-ATPase (green), LPG (red) and DNA (blue) (A) or V-ATPase (green), LAMP-1 (red) and DNA (blue) (B). A and B, Confocal images illustrating V-ATPase acquisition on parasite-containing phagosomes (white arrowheads). C, V-ATPase acquisition was determined on at least 100 phagosomes for each condition and expressed as a percentage of recruitment. Three independent experiments were performed and the bars show the standard deviations of one representative triplicate (**, p=0.005; p values compare the acquisition of V-ATPase on phagosomes containing promastigotes vs amastigotes parasites). Bar, 3 µm.

From: Vinet AF, Fukuda M, Turco SJ, Descoteaux A (2009) The Leishmania donovani Lipophosphoglycan Excludes the Vesicular Proton-ATPase from Phagosomes by Impairing the Recruitment of Synaptotagmin V. PLoS Pathog 5(10): e1000628.

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Leishmania LPG Antibody | CA7AE gallery image 5

Published investigator image:
CsA-treated L. donovani promastigotes show altered morphology. Promastigotes were incubated with 0.15% ethanol or 15 µM (B, C) or 20 µM (A) CsA at 26°C, pH 7.4 for 72 hours. Axenic amastigotes were prepared as described in experimental procedure. 107 cells were fixed with either methanol for Giemsa staining (A), or 2.5% glutaraldehyde for scanning electron microscopy (B). The bar corresponds to 1 µm (B) and 5 µm (A). Two independent experiments were performed and representative fields are shown. (C) Flagellum length measurement. CsA-treated and solvent treated cells L. donovani promastigotes were fixed in methanol and stained with anti-tubulin monoclonal antibody. Flagellum length was measured from a total of 180 cells each for control and CsA-treated samples. Only cells with a single flagellum that was completely visible and fully in focus were taken into account. Samples were observed with a DMR Leica microscope and images were captured with a Cool Snap HQ camera (Roper Scientific). Images were analysed using the IPLab Spectrum 3.9 software (Scanalytics & BD Biosciences) and flagellum length was measured using ImageJ (NIH). (D) Immunoblot analysis of CsA treated parasites. Parasites were treated with solvent or 15 µM CsA for 72 hours, lysed in 1× Laemmli buffer, and lysates equal to 2×107 cells were analyzed by immunoblotting. Promastigote specific marker LPG (upper), amastigote specific marker A2 (middle) and a-tubulin (lower) were analyzed. Two independent experiments which gave identical results were performed.

From: Yau W-L, Blisnick T, Taly J-F, Helmer-Citterich M, Schiene-Fischer C, et al. (2010) Cyclosporin A Treatment of Leishmania donovani Reveals Stage-Specific Functions of Cyclophilins in Parasite Proliferation and Viability. PLoS Negl Trop Dis 4(6): e729.

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  • Mouse anti Leishmania LPG (Repeat Epitope)
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  • Product Type
    Monoclonal Antibody
  • Clone
    CA7AE
  • Isotype
    IgM
1 Formats Available
    Product CodeApplicationsDatasheetMSDSPack SizeList PriceQuantity
    OBT2002E, IB, IFdatasheet pdfdatasheet pdf0.5 ml
    OBT2002
    Summary
    Secondary Antibodies
    Negative Isotype Controls
    Useful Reagents
    Positive Controls
    Histology Controls
    More Images
    References
    Reviews
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    • Mouse anti Leishmania lipophosphoglycan antibody, clone CA7AE recognizes lipophosphoglycan (LPG) the major cell surface glycoconjugate of Leishmania parasites. Mouse anti Leishmania lipophosphoglycan antibody, clone CA7AE recognises the repeat carbohydrate epitope of most species of Leishmania LPG. The epitope is also found on the excreted acid phosphatase of Leishmania and is expressed on the surface of Leishmania infected macrophages (Tolson et al. 1990).

      Clone CA7AE recognizes the promastigotes of Leishmania donovani but not those of the related species L. tropica (Jaffe and Sarfstein 1987, Sundar et al. 2001).
    • Intended Use
    • Target Species
      Protozoan
    • Product Form
      Ascites - lyophilised
    • Reconstitution
      Reconstitute with 0.5 ml distilled water
      Care should be taken during reconstitution as the protein may appear as a film at the bottom of the vial. Bio-Rad recommend that the vial is gently mixed after reconstitution. For long term storage the addition of 0.09% sodium azide is recommended.
    • Preparation
    • Preservative Stabilisers
      None present
    • Immunogen
      Heat killed Leishmania donovani promastigotes.
    • Purity
    • Approx. Protein Concentrations
    • Reagents In The Kit
    • Preparing The Antibody
    • Test Principle
    • Buffer Solution
    • Fusion Partners
      Spleen cells from immunised BALB/c mice were fused with cells of the murine SP2/0 myeloma cell line.
    • Storage
      Prior to reconstitution store at +4oC.
      After reconstitution store at -20oC.
      Storage in frost-free freezers is not recommended. This product should be stored undiluted. Avoid repeated freezing and thawing as this may denature the antibody.
    • Shelf Life
      12 months from date of reconstitution.
    • Acknowledgements
    • Regulatory
      For research purposes only
    • This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit the antibody protocols page.

    • Application NameYesNoMin DilutionMax Dilution
      ELISA1/1000
      Immunoblotting
      Immunofluorescence1/5001/1000

    • Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
    • Technical Advice
    • Recommended Protocol
    • ELISA
    • Immunohistology
    • Histology Positive Control Tissue
    • Immunofluorescence
    • Western Blotting
    • Instructions For Use

    Additional Leishmania LPG Antibody Formats

    Formats Clone Applications Sizes available
    Leishmania LPG Antibody : Ascites CA7AE E, IB, IF 0.5 ml
    • Copyright © 2016 Bio-Rad

    Recommended Secondary Antibody

      Recommended Negative Isotype Control

        Useful Reagents

          Recommended Positive Controls

            Histology Controls

              References

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