“Adherent” versus Other Isolation Strategies for Expanding Purified, Potent, and Activated Human NK Cells for Cancer Immunotherapy

Table 3

Change in the percent and MFI of CD62L on CD3⁻CD56⁺ cells from healthy donor PBMCs after enrichment and expansion. Analysis of CD62L expression on CD3⁻CD56⁺ cells before and after enrichment and expansion. PBMC populations and two-week expanded cells were stained with Pacific Blue conjugated anti-CD3 (Clone HIT3a, Biolegend), Phycoerythrin-Cyanine 7 (PE/Cy7) conjugated anti-CD56 (Clone MEM-188, Biolegend), and Allophycocyanin (APC) conjugated anti-CD62L (Clone P3.6.2.8.1, eBioscience) antibodies and fixed in 2% paraformaldehyde. For all flow cytometry experiments, appropriate IgG isotype controls were used to assess nonspecific staining. Cells were analyzed using a BD LSRII flow cytometer (BD Biosciences) and the data was analyzed using FlowJo Flow Cytometry Analysis Software (TreeStar Inc.).

Donor	In PBMCs			In expanded NK cells on day 14			Culture-induced change
	Percent NK cells	Percent CD62L^+ NK cells	CD62L MFI of NK cells	Percent NK cell purity	Percent CD62L^+ NK cells	MFI of CD62L	Change in CD62L^+ NK cells after expansion	Change in CD62L^+ NK cell MFI after expansion
2	13.1	30.8	5,647	94.2	24.8	4,383	19.5% lower	22.4% lower
JL	17.2	17.3	5,689	84.8	35.3	4,394	104.0% higher	32.8% lower
4	5.7	14.4	2,846	69.8	25.7	3,579	78.5% higher	25.8% higher
AS	17.0	68.5	13,316	95.2	62.9	9,119	8.2% lower	31.5% lower
SS	15.7	69.0	6,672	97.9	34.9	3,373	49.4% lower	49.4% lower
^*SS	13.9	65.7	6,927	98.8	50.1	5,891	23.7% lower	15.0% lower
21	19.8	36.0	5,891	94.9	14.7	2,953	59.2% lower	49.1% lower
^*21	19.8	36.0	5,891	92.0	23.6	2,537	34.4% lower	56.9% lower

Large-scale expansion.