Precision and linearity targets for validation of an IFNγ ELISPOT, cytokine flow cytometry, and tetramer assay using CMV peptides

Table 1 Mean response levels of the three CMV-responsive donors

	% CD3⁺CD8⁺ cells	ELISPOT (peptide mix)	CFC (peptide mix)	CFC (pp65_495–503)	Tetramer (pp65_495–503)	ELISPOT (pp65_495–503)
Donor 41 (low)	20.3%	270 SFC¹ [0.53%]	nd²	0.06%²	0.06%²	8 SFC¹ [0.02%]
Donor 68 (med)	12.1%	285 SFC¹ [0.94%]	0.28%²	0.28%²	0.25%²	31 SFC¹ [0.10%]
Donor 43 (high)	26.3%	95 SFC¹ [0.14%]	0.87%²	0.93%²	1.27%²	28 SFC¹ [0.04%]

¹ ELISPOT results reported as SFC per 2.5 × 10⁵ PBMC. Numbers in brackets indicate the values when re-calculated as % of CD8⁺ T cells. Note that ELISPOT responses with peptide mix did not match the other assays in terms of low, medium, and high responders.
² CFC and tetramer results reported as % of CD8⁺ T cells. nd = CFC was not done on this donor due to predicted very low response (0.02%) seen in pre-screening.

ISSN: 1471-2172