ATLA::Alternatives to Laboratory Animals

Volume 30, Number 4

Stable biocompatible adjuvants -- a new type of adjuvant based on solid lipid nanoparticles: a study on cytotoxicity, compatibility and efficacy in chicken

Carsten Olbrich,l Rainer Helmut Müller,1 Kerstin Tabatt,l Oliver Kayser,1 Christoph Schulze2 and Rüdiger Schade3

1Department of Pharmaceutical Technology, Biopharmacy and Biotechnology, Free University of Berlin, Kelchstrasse 31, 12169 Berlin, Germany; 2State Department for Consumer Protection, Ringstrasse 1030, 15234 FrankfurtlOder, Germany; 3Institute of Pharmacology and Toxicology, Medical Faculty (Charité), Humboldt-University, Dorotheenstrasse 94, 10117 Berlin, Germany

SUMMARY

A new type of adjuvant was tested for its ability to initiate antibody production in chickens, and its cellular and tissue compatibility were assessed. The stable biocompatible adjuvants tested are based on surface-modified solid lipid nanoparticles (SLNs), made from paraffin or biodegradable glycerides, and are simply admixed to the antigens before administration. The tissue-damaging potency of four formulations of the new adjuvants (H1, H2, H3 and H4) were first tested in vitro by using human foreskin fibroblasts and RAW 264.7 macrophages. The adjuvants were well tolerated by both cell types. Immunisation studies in chickens were performed by using a Mycoplasma bovis antigen and mouse immunoglobulin G (IgG). The resulting antibodies were non-invasively extracted from egg yolk. The use of the various adjuvant formulations resulted in a significant production of specific antibodies after the first and second booster immunisations. Freund's complete adjuvant (FCA), considered until now to be the "gold standard" among the adjuvants, revealed the highest antibody titre against mouse IgG. SLNs with a particle size of more than 100 nm exhibited a clear adjuvant activity, whereas SLNs with a particle size below 100 nm, in various concentrations, revealed a lower adjuvant activity. Immunisation of chickens with the mouse IgG alone, dissolved in phosphate-buffered saline, resulted in a slow development of antibody titre. At the end of the experiment, the chickens were examined for vaccination-associated tissue damage. In contrast to FCA, the SLN formulations caused only minor tissue irritation at the injection sites. In conclusion, SLNs seem to be a promising alternative to FCA for antibody production in chickens, and potentially in other animals.

Keywords: chicken antibodies, compatibility, IgY, new adjuvant, solid lipid nanoparticles