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. 2001 Oct;126(1):165–172. doi: 10.1046/j.1365-2249.2001.01622.x

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© 2001 Blackwell Science Ltd

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Fig. 7 — FACS, Northern blotting and NK cell-mediated cytotoxicity assay of 721.221 transfectants with HLA-G3. (a) Northern blotting of the transfectants is shown. The probe used for the hybridization was the cDNA restriction fragment of HLA-G α1 domain. (b) NK cell-mediated cytotoxicity assay. The parental 721.221 cell or transfected cells were incubated with PBMC at the E:T ratio 30:1. The results are expressed as a percentage of specific lysis. Values are presented as mean ± s.e.m. (n = 6). (□) Naive 721.221; (▪) 721.221-G3-1; (▪) 721.221-G3-2; () 721.221-mock. (c) FACS profiles of 721.221 transfectants with HLA-G3. Naive cell (dotted line) and 721.221 transfectants, 721.221-G3-2, were treated with anti-HLA class I MoAb, B9–12–1 (thick line), or control isotype antibody (thin line). (d) Blocking assay by anti-CD94 MoAb. To determine whether the down-regulatory effect is related to the CD94/NKG2 receptor, NK cell-mediated cytotoxicity assay was performed using HLA-G3 transfected 721.221 cells, 721.221 – G3–2, and mock transfectant, 721.221-mock, as the control. An IgG2a isotype MoAb was also used as a control MoAb. Relative lysis of 721.221 – G3–2 with or without anti-CD94 MoAb is shown compared with the mock transfectant cell line. Values are presented as mean ± s.e.m/ (n = 6). Indicates significant difference (P < 0·05). (□) No antibody; (░) control antibody; (▪) anti-CD94 antibody.

Inline graphic — FACS, Northern blotting and NK cell-mediated cytotoxicity assay of 721.221 transfectants with HLA-G3. (a) Northern blotting of the transfectants is shown. The probe used for the hybridization was the cDNA restriction fragment of HLA-G α1 domain. (b) NK cell-mediated cytotoxicity assay. The parental 721.221 cell or transfected cells were incubated with PBMC at the E:T ratio 30:1. The results are expressed as a percentage of specific lysis. Values are presented as mean ± s.e.m. (n = 6). (□) Naive 721.221; (▪) 721.221-G3-1; (▪) 721.221-G3-2; () 721.221-mock. (c) FACS profiles of 721.221 transfectants with HLA-G3. Naive cell (dotted line) and 721.221 transfectants, 721.221-G3-2, were treated with anti-HLA class I MoAb, B9–12–1 (thick line), or control isotype antibody (thin line). (d) Blocking assay by anti-CD94 MoAb. To determine whether the down-regulatory effect is related to the CD94/NKG2 receptor, NK cell-mediated cytotoxicity assay was performed using HLA-G3 transfected 721.221 cells, 721.221 – G3–2, and mock transfectant, 721.221-mock, as the control. An IgG2a isotype MoAb was also used as a control MoAb. Relative lysis of 721.221 – G3–2 with or without anti-CD94 MoAb is shown compared with the mock transfectant cell line. Values are presented as mean ± s.e.m/ (n = 6). Indicates significant difference (P < 0·05). (□) No antibody; (░) control antibody; (▪) anti-CD94 antibody.