Table 3. Polyclonal and mono/oligoclonal... | American Society of Hematology

Table 3.

Polyclonal and mono/oligoclonal reconstitution of NOD/SCID mice by lentivirally transduced SRC

Limiting dilution	Primary recipients						Secondary recipients
Group	1° mouse	Cell dose	CD45⁺FACS (%)	GFP⁺ FACS (%)	Multilineage engraftment	2° CFU-GM GFP (%)	2° mouse	CD45⁺FACS (%)	GFP⁺ FACS (%)	Multilineage engraftment	3° CFU-GM GFP (%)
V	5.1	400 000	73	14	yes	10	5.1.1	0.05	0.5	no	NA
	5.2		22	11	yes	ND	5.1.2	0.04	0	no	NA
	5.3		28	29	yes	30	5.1.3	dead			NA
	5.4		36	14	yes	ND
							5.2.1	0.05	10	no	NA
VI	6.5	100 000	1.8	1.6	yes	ND	5.2.2	0.02	0	no	NA
	6.6		6.7	21	yes	0	5.2.3	dead			NA
	6.7		5.5	22	yes	ND
	6.8		22	9.3	yes	ND	5.3.1	0.05	0	no	NA
							5.3.2	0.5	5	yes	0
VII	7.21	35 000	0.28	52	yes	100	5.3.3	0.1	64	yes	74
	7.22		21	76	yes	67
	7.23		3.8	91	yes	100	5.4.1	0.04	0	no	NA
	7.24		0.01	0	no	NA	5.4.2	0.03	0	no	NA
							5.4.3	dead			NA
VIII	8.25	12 500	4.2	16	yes	13
	8.26		0.52	57	yes	0	6.5.1	0.03	0	no	NA
	8.27		0.01	0	no	NA	6.6.1	0.2	22	no	NA
	8.28		0	0	no	NA	6.7.1	0.05	0	no	NA
IX	9.29	5000	1.2	0	no	NA	6.8.1	0.2	0	no	NA
	9.30		3.2	0	no	NA
	9.31		0.11	0	no	NA
	9.32		0.01	0	no	NA

Limiting dilution	Primary recipients						Secondary recipients
Group	1° mouse	Cell dose	CD45⁺FACS (%)	GFP⁺ FACS (%)	Multilineage engraftment	2° CFU-GM GFP (%)	2° mouse	CD45⁺FACS (%)	GFP⁺ FACS (%)	Multilineage engraftment	3° CFU-GM GFP (%)
V	5.1	400 000	73	14	yes	10	5.1.1	0.05	0.5	no	NA
	5.2		22	11	yes	ND	5.1.2	0.04	0	no	NA
	5.3		28	29	yes	30	5.1.3	dead			NA
	5.4		36	14	yes	ND
							5.2.1	0.05	10	no	NA
VI	6.5	100 000	1.8	1.6	yes	ND	5.2.2	0.02	0	no	NA
	6.6		6.7	21	yes	0	5.2.3	dead			NA
	6.7		5.5	22	yes	ND
	6.8		22	9.3	yes	ND	5.3.1	0.05	0	no	NA
							5.3.2	0.5	5	yes	0
VII	7.21	35 000	0.28	52	yes	100	5.3.3	0.1	64	yes	74
	7.22		21	76	yes	67
	7.23		3.8	91	yes	100	5.4.1	0.04	0	no	NA
	7.24		0.01	0	no	NA	5.4.2	0.03	0	no	NA
							5.4.3	dead			NA
VIII	8.25	12 500	4.2	16	yes	13
	8.26		0.52	57	yes	0	6.5.1	0.03	0	no	NA
	8.27		0.01	0	no	NA	6.6.1	0.2	22	no	NA
	8.28		0	0	no	NA	6.7.1	0.05	0	no	NA
IX	9.29	5000	1.2	0	no	NA	6.8.1	0.2	0	no	NA
	9.30		3.2	0	no	NA
	9.31		0.11	0	no	NA
	9.32		0.01	0	no	NA

NA indicates not applicable due to absence of human CFU; ND, not done.

A cell-dose–limiting dilution of transduced CD34⁺ cells (MOI = 200) transplanted into NOD/SCID recipients shows successful lentiviral vector gene transfer into both primary and secondary SRC. Bone marrow from high-cell-dose primary transplant recipients (groups V and VI) were transplanted into secondary recipients with one donor for one recipient mouse for group VI donors and one donor for three recipient mice for group V donors. FACS analysis for engraftment (CD45) and GFP expression was performed along with colony analysis for GFP expression by microscopy. Multilineage engraftment was seen as defined by the presence of CFU-GM and a minimum of 5 human lymphoid cells (CD19) and 5 human myeloid cells (CD15 or CD33) per 20 000 mouse bone marrow mononuclear cells.

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