TABLE 5.

Detection of antigen-specific cells by ELISPOT^a

Donor	Titer
	Medium	PHA	SEA	Tetanus toxoid	Heat-inactivated HIVSX	Mock infected
HIV seropositive
60016b	<1/200,000	1/1,471	NDe	1/28,571	1/33,333	ND
60030c	<1/200,000	ND	1/2,595	1/23,529	1/26,667	<1/200,000
60031d	<1/200,000	ND	1/1,235	1/10,526	1/12,903	<1/200,000
HIV seronegative
60001	<1/200,000	ND	1/2,702	1/31,746	<1/200,000	<1/200,000
60002	<1/200,000	ND	1/2,381	1/12,500	<1/200,000	<1/200,000

^aDuplicate wells of DC were cultured in serum-free medium supplemented with 1,000 U each of IL-4 and GM-CSF per ml and 10 U of IFN-γ per ml for 4 days. On day 4, DC were pulsed with tetanus toxoid (2 μg/ml), SEA (1 μg/ml), heat-inactivated (for 30 min at 62°C) HIV_SX (40 ng/ml), or supernatants derived from mock-infected cultures from the same donors in whom HIV_SX was grown (volume equal to that for heat-inactivated HIV) for 16 h. After 16 h, the Ags and cytokines were removed and 2 × 10⁵ autologous PBMC were added to the DC for 6 h. The PBMC were then transferred to 96-well PVDF-backed plates (Millipore) coated with anti-IFN-γ MAb 1-D1K (Mabtech) and were incubated at 37°C for 48 h. The ELISPOT assay was performed as instructed by the manufacturer (Mabtech). All HIV-infected donors were on antiretroviral therapy for at least 1 year. 

^bViral load, <400 copies/ml; CD4⁺ T cells, 540/mm³. 

^cViral load, <400 copies/ml; and CD4⁺ T cells, 800/mm³. 

^dViral load, <40 copies/ml; CD4⁺ T cells, 921/mm³. 

^eND, not determined.