Borrelia Burgdorferi antibody

Catalog No. ABIN236885

Product Details anti-Borrelia Burgdorferi Antibody

Target: Borrelia Burgdorferi

Reactivity: Borrelia burgdorferi

Host: Rabbit

Clonality: Polyclonal

Conjugate: This Borrelia Burgdorferi antibody is un-conjugated

Application: Western Blotting (WB), Immunofluorescence (IF), Immunohistochemistry (Paraffin-embedded Sections) (IHC (p))

Specificity: Reactive Western blot bands at 83kD, 41kD, 34kD and 31kD and additional low MW bands. Cross-reacts with Treponema pallidum, B. hermsii and B. parkerii.

Characteristics: Rabbit anti B. burgdorferi, Rabbit antibody to Borrelia burgdorferi

Purification: Protein A Chromatography. Ascites

Purity: > 90 % pure

Immunogen: Whole Cell preparation from B. burgdorferi Strain: B31 ATCC 35210

Application Details

Application Notes: Suitable for use in IFA, Western blot and immunohistochemistry (formalin fixed/paraffin). IFA specimens should be acetone-fixed for best results. Also suitable for conjugation purposes. Each laboratory should determine an optimum working titer for use in its particular application. Other applications have not been tested but use in such assays should not necessarily be excluded.

Restrictions: For Research Use only

Independent Validation (IF)

Validation #100172 (Immunofluorescence)

by: Institut für Medizinische Mikrobiologie, Virologie und Hygiene, Universitätsklinikum Hamburg-Eppendorf

No.: #100172

Date: 01/18/2017

Antigen: Borrelia Burgdorferi

Lot Number: 10L35615

Method validated: Immunofluorescence

Positive Control: Macrophages expressing a protein X locating at plasma and endomembranes, infected with Borrelia burgdorferi B31.

Negative Control: Macrophages expressing a protein X locating at plasma and endomembranes.

Notes: Passed. ABIN236885 has been shown to specifically label Borrelia burgdorferi B31 inside human macrophages and extracellularly.

Validation Images

Staining of a human macrophage after infection with Borrelia burgdorferi B31. A. shows labeling of a protein associated to plasma membranes and endomembranes. B. and C. depict labeling of Borrelia burgdorferi B31 inside and outside of the cell respectively with ABIN236885. Panel D. is an overlay of all three channels.

Full Methods

Primary Antibody: ABIN236885

Secondary Antibody: anti-rabbit-AF488 (Jackson ImmunoResearch Inc, 111-545-003), anti-rabbit-Alexa568 (Thermo Fisher Scientific, A11011)

Full Protocol:

• Transfection and infection:
• • Isolate human peripheral blood monocytes from buffy coats and differentiate them into macrophages in RPMI growth media with 20% human serum and 5% L-Glutamin-Pen/Strep (Sigma-Aldrich, G6784) at 37°C, 5% CO₂.
  • Transfect primary human macrophages with an expressing plasmid for a protein X that binds to plasma membranes and endomembranes using Neon Transfection System (Thermo Fisher Scientific, MPK5000) for 3h at 37°C and seeded on 12mm glass coverslips (see http://www.ncbi.nlm.nih.gov/pubmed/24696301).
  • Infect macrophages expressing protein X with Borrelia burgdorferi B31 for 1h at 37°C and MOI 100.
  • Fix cells 10 min with 4% methanol-free formaldehyde.
• Extracellular immunofluorescence staining.
• • Block with 40µl of 2% BSA/PBS containing 5% normal human serum (Jackson ImmunoResearch Inc, 009-000-001) and 5% normal goat serum (Jackson ImmunoResearch Inc, 005-000-001) for 30min at RT.
  • Wash 2x with PBS.
  • Incubation with 40µl primary rabbit-anti-Borrelia burgdorferi antibody (antibodies-online, ABIN236885, lot 10L35615) diluted 1:1000 in PBS for 30min at RT.
  • Wash 2x with PBS.
  • Incubation with 40µl secondary anti-rabbit-AF488 antibody (Jackson ImmunoResearch Inc, 111-545-003) diluted 1:200 in PBS for 30min at RT.
  • Wash 2x with PBS.
• Intracellular immunofluorescence staining:
• • Permeabilize cells with 0.05% Triton X-100/PBS for 10 min at RT.
  • Wash 2x with PBS.
  • Incubation with 40µl primary rabbit-anti-Borrelia burgdorferi antibody (antibodies-online, ABIN236885, lot 10L35615) diluted 1:1000 in PBS for 30min at RT.
  • Wash 2x with PBS.
  • Incubation with 40µl of anti-rabbit-AF568 antibody (Thermo Fisher Scientific, A11011) diluted 1:200 for 30 min at RT.
  • Wash 2x with PBS.
  • Mount coverslips with 6µl prewarmed Mowiol 4-88 mounting medium (Sigma-Aldrich, 81381) on object slides for at least 1h.
• Image acquisition:
• • Image acquisition was performed with a Leica TCS SP5 (with a Leica TCS SP5 AOBS confocal point scanner), equipped with an oil-immersion HCX PL APO 63x NA 1.4 λ_blue objective.

Handling

Format: Liquid

Concentration: 4-5 mg/mL

Buffer: 0.01 M PBS, pH 7.2 Product contains no stabilizing proteins, 0.1 % Sodium azide

Preservative: Sodium azide

Precaution of Use: This product contains sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.

Handling Advice: Avoid multiple freeze/thaw cycles.
Centrifuge product if not completely clear after standing at room temperature.
Prepare working dilution only prior to immediate use.

Storage: 4 °C/-20 °C

Storage Comment: Short-term (up to 6 months) store at 2-8 °C. Long term, aliquot and store at -28 °C.

Expiry Date: 6 months

References for anti-Borrelia Burgdorferi antibody (ABIN236885)

Klose, Scheungrab, Luckner, Wanner, Linder: "FIB-SEM-based analysis of Borrelia intracellular processing by human macrophages." in: Journal of cell science, Vol. 134, Issue 5, (2021) (PubMed).

Naj, Linder: "ER-Coordinated Activities of Rab22a and Rab5a Drive Phagosomal Compaction and Intracellular Processing of Borrelia burgdorferi by Macrophages." in: Cell reports, Vol. 12, Issue 11, pp. 1816-30, (2015) (PubMed).

Hoffmann, Naj, Linder: "Daam1 is a regulator of filopodia formation and phagocytic uptake of Borrelia burgdorferi by primary human macrophages." in: FASEB journal : official publication of the Federation of American Societies for Experimental Biology, Vol. 28, Issue 7, pp. 3075-89, (2014) (PubMed).

Target Details for Borrelia Burgdorferi

Target: Borrelia Burgdorferi

Abstract: Borrelia Burgdorferi Products