Assessment of anti-bovine IL4 and IFN gamma antibodies to label IL4 and IFN gamma in lymphocytes of the koala and brushtail possum
Damien P. Higgins*, Susan Hemsley, Paul J. Canﬁeld
Faculty of Veterinary Science, University of Sydney, Sydney, NSW 2006, Australia
We assess anti-bovine IL4 and IFN gamma (IFNg) antibodies for their ability to label IL4 and IFNg in koala (Phascolarctos cinereus), common brushtail possum (Trichosurus vulpecula) and mountain brushtail possum (Trichosurus caninus) lymphocytesusing ﬂow cytometry and immunohistochemistryto determine their applicability to studies of host response to intracellular pathogens. Anti-IFNg labelled a product of PMA-ionomycin stimulated sheep, koala and possum lymphocytes. High intensity labelling was not reduced by blocking non-speciﬁc binding with 10% FCS; and non-permeabilised koala lymphocytes labelled less, demonstrating that the labelled product was intracellular. The anti-IL4 antibody labelled variably more cells than the irrelevant antibodyin some stimulated and non-stimulated preparations in all species but intensity of this labelling was similar to that of cells labelled with the irrelevant antibody. In this study, the antibodies did not label frozen or formalin-ﬁxed tissues in a range of immunohistochemical techniques. We expect the anti-IFNg antibody to be effective in evaluating Th1 responses of koalas and possums exposed to various host, pathogen and environmental factors and add to the limited tools available for investigating the pathogenesis of marsupial diseases, especially those caused by intracellular organisms, such as tuberculosis of brushtail possums and chlamydial disease of koalas. # 2004 Elsevier B.V. All rights reserved.