Ee. Doofin et al., Purification of secretory immunoglobulin A from milk of the brushtail possum (Trichosurus vulpecula), NZ VET J, 49(5), 2001, pp. 181-186
AIM: To identify and purify secretory immunoglobulin A (sIgA), a key effect
er molecule in mucosal immune responses, from milk of the brushtail possum
(Trichosurus vulpecula).
METHODS: Milk samples were collected from female possums with pouch young,
and clarified by centrifugation and precipitation methods. The clarified fr
action was purified by gel filtration and affinity chromatography to yield
sIgA. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE)
and immunoblotting techniques were used to assess the purity of the final
product, and to identify the heavy (H) chain, light (L) chain and secretory
component (SC) of possum sIgA.
RESULTS: Immunoblotting, using antibodies raised against cloned possum sIgA
SC and H-chain, and a synthetic peptide fragment of the H-chain, confirmed
the identity of the purified protein. The N-terminal amino acid sequence o
f purified possum sIgA showed strong homology to reported sequences of H-ch
ain variable regions of marsupial immunoglobulins.
CONCLUSIONS: Milk was shown to be a convenient source of mucosal secretion
containing sIgA, and a process involving 2 precipitation and 2 chromatograp
hy steps produced purified sIgA. This IgA preparation will prove useful for
the generation of sIgA-specific immunological reagents for measurement of
immune responses in the development of mucosal-based vaccines for biologica
l control of possums.