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Antibodies 2013, 2(1), 66-81; doi:10.3390/antib2010066

Shark Variable New Antigen Receptor (VNAR) Single Domain Antibody Fragments: Stability and Diagnostic Applications

1
The Department of Biochemistry, La Trobe Institute for Molecular Science, La Trobe University, Plenty Road, Bundoora, VIC., 3086, Australia
2
AdAlta Pty. Ltd., 15/2 Park Drive, Bundoora, VIC., 3086, Australia
3
The CSIRO Materials Science and Engineering, 343 Royal Parade, Parkville, VIC., 3052, Australia
4
The Foundation for Innovative New Diagnostics (FIND), Avenue de Budé 16, 1202 Geneva, Switzerland
*
Author to whom correspondence should be addressed.
Received: 29 November 2012 / Revised: 9 January 2013 / Accepted: 21 January 2013 / Published: 25 January 2013
(This article belongs to the Special Issue Single-Domain Antibody)
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Abstract

The single variable new antigen receptor domain antibody fragments (VNARs) derived from shark immunoglobulin new antigen receptor antibodies (IgNARs) represent some of the smallest known immunoglobulin-based protein scaffolds. As single domains, they demonstrate favorable size and cryptic epitope recognition properties, making them attractive in diagnosis and therapy of numerous disease states. Here, we examine the stability of VNAR domains with a focus on a family of VNARs specific for apical membrane antigen 1 (AMA-1) from Plasmodium falciparum. The VNARs are compared to traditional monoclonal antibodies (mAbs) in liquid, lyophilized and immobilized nitrocellulose formats. When maintained in various formats at 45 °C, VNARs have improved stability compared to mAbs for periods of up to four weeks. Using circular dichroism spectroscopy we demonstrate that VNAR domains are able to refold following heating to 80 °C. We also demonstrate that VNAR domains are stable during incubation under potential in vivo conditions such as stomach acid, but not to the protease rich environment of murine stomach scrapings. Taken together, our results demonstrate the suitability of shark VNAR domains for various diagnostic platforms and related applications.
Keywords: single-domain antibody; shark VNAR; thermal stability; pH stability; protease resistance; AMA-1; Plasmodium; malaria; diagnosis single-domain antibody; shark VNAR; thermal stability; pH stability; protease resistance; AMA-1; Plasmodium; malaria; diagnosis
This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).

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MDPI and ACS Style

Griffiths, K.; Dolezal, O.; Parisi, K.; Angerosa, J.; Dogovski, C.; Barraclough, M.; Sanalla, A.; Casey, J.L.; González, I.; Perugini, M.A.; Nuttall, S.; Foley, M. Shark Variable New Antigen Receptor (VNAR) Single Domain Antibody Fragments: Stability and Diagnostic Applications. Antibodies 2013, 2, 66-81.

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