Replicase-based DNA vaccines for allergy treatment.

Replicase-based vaccines were introduced to overcome some of the deficiencies of conventional DNA- and RNA-based vaccines, including poor efficiency and low stability. At ultra-low doses, these alphavirus-derived vectors elicit cellular as well as humoral immune responses. Additionally, replicase-based vectors induce "self-removal" of the vaccine via apoptosis of transfected cells. This chapter describes the construction of a replicon-based DNA vaccine vector from commercially available plasmids. We present protocols for monitoring cellular immune responses following replicase-based immunization including measurement of allergen-specific proliferation of splenocytes, ELISPOT, a FACS-based cytokine secretion assay providing information about T-helper subsets, and a cytokine fluorescent bead immunoassay.