This Article Full Text Full Text (PDF) Submit a response Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this link to a friend Similar articles in this journal Similar articles in PubMed Add article to my folders Download to citation manager Request Permissions Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Brereton, H M Articles by Williams, K A Search for Related Content PubMed PubMed Citation Articles by Brereton, H M Articles by Williams, K A

Influence of format on in vitro penetration of antibody fragments through porcine cornea

¹ Department of Ophthalmology, Flinders University, Adelaide, Australia
² CSL Ltd, Parkville, Victoria, Australia
³ Department of Ophthalmology, University of Zurich, Zurich, Switzerland

Correspondence to:
Correspondence to:
Dr Keryn Williams
Department of Ophthalmology, Flinders Medical Centre, Bedford Park, SA 5042, Australia; keryn.williams{at}flinders.edu.au

Aim: Antibody fragments, appropriately formulated, can penetrate through the ocular surface and thus have potential as therapeutic agents. The aim was to investigate the influence of protein fragment format on the kinetics and extent of ocular penetration in vitro.

Methods: Immunoglobulin single chain variable domain fragments of a murine monoclonal antibody with specificity for rat CD4 were engineered with a 20 or 11 amino acid linker by assembly polymerase chain reaction, expressed in Escherichia coli and purified by chromatography. Fab fragments of the parental antibody were prepared by papain digestion. Antibody fragments were formulated with a penetration and a viscosity enhancer and were applied to the surface of perfused pig corneas for up to 10 hours in vitro. Penetration was quantified by flow cytometry on rat thymocytes.

Results: 20-mer antibody fragments formed natural monomers and dimers following purification that could be separately isolated, while 11-mer fragments were dimeric. All formats of fragment (20-mer monomers and dimers, 11-mer dimers, Fab) showed penetration through the pig cornea after 6 hours of intermittent topical administration.

Conclusion: Antibody fragments of different shapes and sizes can penetrate the cornea after topical administration, thereby increasing the potential of this class of proteins for topical ophthalmic use.

Abbreviations: Ig, immunoglobulins; IMAC, immobilised metal affinity chromatography; MFI, mean fluorescence intensity; scFv, single chain variable domain antibody fragments

Keywords: antibody fragments; cornea; perfusion chamber; drug delivery; pig model