Abstract
The rodent parasite Nippostrongylus brasiliensis (N. brasiliensis) models the salient features of helminth infection including skin penetration, migration from tissues to lung, maturation and egg production in the gut. As a potent activator of systemic and mucosal Th2 immune responses, Nippostrongylus brasiliensis has been extensively used to study host protective immunity and in vivo regulation of Th2 immune response. Six to eight week old C57Bl/6J, Balb/c mice or any other strains are suitable, as all are susceptible to infection. Inocula of 150-650 L3 larvae can be administered by subcutaneous injection, but for greatest consistency a dose of 550 L3 larvae is routinely used for experimental purposes. We have optimized three different protocols for the isolation of larvae from the lungs of mice infected with the L3 stage of Nippostrongylus brasiliensis. Larvae can migrate to the lung between 18-60 h post inoculation from any site in the body. The numbers of larvae appearing in the lung peaks at 48 h after inoculation and it is recommended that isolation/harvesting be performed at 48 h for greatest consistency of each harvest method:
Part I. Isolation by migration
Materials and Reagents
Equipment
Procedure
Part II. Isolation by lung digest
Part III. Isolation by bronchoalveolar lavage (BAL) This protocol has been adapted from protocols used in measuring allergic airway inflammation (Harris et al., 1997; Erb et al., 1998).
Representative data
Figure 4. Comparison of larval recovery from lung using either thermal gradient induced migration, bronchoalveolar lavage (BAL) or lung digestion
Notes
For consistent results using the above protocols, it is important to always use larvae from 2-6 weeks old cultures for infecting mice. The infectivity of larvae is much reduced after 6 weeks culture (Camberis et al., 2003; Kassai, 1982).
Recipes
Acknowledgments
This work has been adapted and modified from previous work carried out by Professor Graham Le Gros’s Allergy and Parasitic diseases laboratory, Malaghan Institute of Medical Research, New Zealand. This work was supported by The Health Research Council of New Zealand and the Marjorie Barclay Trust.
References
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