Development of a limiting dilution assay to enumerate the frequency of Helper T Lymphocyte precursor cells specific for Herpes Simplex Virus Type 1

A limiting dilution assay system was developed to quantitate the frequency of Helper T Lymphocyte-precursors (HTL-P) specific for Herpes Simplex Virus Type 1 (HSV-1). Limiting dilution assay conditions for the estimation of optimal helper cell frequencies were determined. The HSV-specific HTL-P was demonstrated to be virus-specific for in vivo induction, in vitro clonal expansion and in vitro restimulation to promote interleukin 2 (IL 2) production. The cell type of the HTL-P was shown to be a Thy 1.2⁺, Lyt 1.1⁺ cell and the estimated frequency of the HTL-P could be increased 3-fold by Lyt 2.1 depletion of the responder population.

The utility of this limiting dilution approach in the evaluation of various immunization protocols was demonstrated. Live HSV-1 in vivo priming was shown to be required to stimulate virus-specific HTL-P as both heat- and UV-inactivated virus preparations failed to stimulate detectable helper cell frequencies. Soluble infected-cell extracts containing HSV-1 glycoproteins also failed to induce detectable virus-specific HTL-P.