A Broad Set of Different Llama Antibodies Specific for a 16 KDa Heat Shock Protein of Mycobacterium Tuberculosis

Overview

Journal PLoS One

Specialties General Medicine
Science

Date 2011 Nov 3

PMID 22046343

Citations 7

Authors

Anke K Trilling

Hans de Ronde

Linda Noteboom

Adele van Houwelingen

Margriet Roelse

Saurabh K Srivastava

Willem Haasnoot

Maarten A Jongsma

Arend Kolk

Han Zuilhof

Jules Beekwilder

Affiliations

Soon will be listed here.

Abstract

Background: Recombinant antibodies are powerful tools in engineering of novel diagnostics. Due to the small size and stable nature of llama antibody domains selected antibodies can serve as a detection reagent in multiplexed and sensitive assays for M. tuberculosis.

Methodology/principal Findings: Antibodies for Mycobacterium tuberculosis (M. tb) recognition were raised in Alpaca, and, by phage display, recombinant variable domains of heavy-chain antibodies (VHH) binding to M. tuberculosis antigens were isolated. Two phage display selection strategies were followed: one direct selection using semi-purified protein antigen, and a depletion strategy with lysates, aiming to avoid cross-reaction to other mycobacteria. Both panning methods selected a set of binders with widely differing complementarity determining regions. Selected recombinant VHHs were produced in E. coli and shown to bind immobilized lysate in direct Enzymelinked Immunosorbent Assay (ELISA) tests and soluble antigen by surface plasmon resonance (SPR) analysis. All tested VHHs were specific for tuberculosis-causing mycobacteria (M. tuberculosis, M. bovis) and exclusively recognized an immunodominant 16 kDa heat shock protein (hsp). The highest affinity VHH had a dissociation constant (KD) of 4 × 10(-10) M.

Conclusions/significance: A broad set of different llama antibodies specific for 16 kDa heat shock protein of M. tuberculosis is available. This protein is highly stable and abundant in M. tuberculosis. The VHH that detect this protein are applied in a robust SPR sensor for identification of tuberculosis-causing mycobacteria.

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16 kDa heat shock protein from heat-inactivated Mycobacterium tuberculosis is a homodimer - suitability for diagnostic applications with specific llama VHH monoclonals.

Srivastava S, Ruigrok V, Thompson N, Trilling A, Heck A, van Rijn C PLoS One. 2013; 8(5):e64040.

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