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. 2014 Dec 16;12:336. doi: 10.1186/s12967-014-0336-5

Table 5.

Outcome of CMVpp65-specific T-cell separation by MiniMACS CSA

A 1. Validation run
Positive control CMVpp65 pp Negative control
OF TCF OF TCF OF TCF
volume [ml] 0.6 1.0 0.6 1.0 0.6 1.0
viability [%] 98.81 33.19 98.70 15.63 98.67 12.21
WBCs(CD45+) [x106/ml] 42.70 0.07 47.50 0.01 39.50 0.00
WBCsabs [x106] 25.62 0.07 28.50 0.01 23.70 0.00
T cells(CD3+CD56-) [% of WBCs] 27.26 35.72 27.56 43.33 31.66 15.62
T cells [/μl] 11650.00 24.90 13100.00 6.23 12500.00 0.70
T cellsabs [x106] 6.99 0.02 7.86 0.01 7.50 0.00
T cells(CD3+CD4+) [% of CD3+] 50.34 33.51 50.25 25.85 49.05 63.35
T cells(CD3+CD8+) [% of CD3+] 49.67 66.51 49.77 74.15 50.97 36.71
IFN-γ + T cells [% of CD3 + ] 0.69 82.33 0.23 69.27 0.01 2.89
IFN-γ+ T cells [/μl] 80.33 20.50 30.17 4.32 1.25 0.02
IFN-γ + T cells [x10 4 ] 4.74 1.21 1.79 0.15 0.07 0.00
IFN-γ− T cells [% of CD3+] 99.31 17.67 99.77 30.73 99.99 97.11
IFN-γ− T cells [/μl] 11566.67 4.40 13066.67 1.92 12500.00 0.68
IFN-γ− T cells [x104] 682 026 778 0.07 724 0.02
IFN-γ+ T cells(CD3+) [% of CD4] 0.18 23.44 0.04 9.06 0.01 1.72
IFN-γ + T cells (CD3+CD4+) [% of CD4] 0.37 70.45 0.09 35.32 0.01 2.82
IFN-γ+ T cells(CD3+CD4+) [/μl] 21.67 5.88 5.93 0.57 0.61 0.01
IFN-γ + T cells abs (CD3+CD4+) [x10 4 ] 1.28 0.35 0.35 0.02 0.04 0.00
IFN-γ+ T cells(CD3+) [% of CD8] 0.49 58.79 0.18 59.14 0.00 0.52
IFN-γ + T cells (CD3+CD8+) [% of CD8] 0.95 87.52 0.37 78.94 0.00 1.42
IFN-γ+ T cells(CD3+CD8+) [/μl] 55.00 14.50 24.17 3.65 0.00 0.00
IFN-γ + T cells abs (CD3+CD8+) [x10 4 ] 3.24 0.86 1.44 0.13 0.00 0.01
B 2. Validation run
Positive control CMVpp65 pp Negative control
OF TCF OF TCF OF TCF
volume [ml] 0.6 1.0 0.6 1.0 0.6 1.0
viability [%] 98.72 89.04 98.92 85.16 98.87 29.95
WBCs(CD45+) [x106/ml] 2.38 0.18 2.74 0.00 2.78 0.00
WBCsabs [x106] 1.43 0.18 1.64 0.00 1.67 0.00
T cells(CD3+CD56-) [% of WBCs] 95.94 88.43 89.37 68.87 88.83 55.91
T cells [/μl] 2283.33 162.00 2450.00 1.42 2466.67 11.10
T cellsabs [x106] 1.37 0.16 1.47 0.00 1.48 0.00
T cells(CD3+CD4+) [% of CD3+] 71.87 75.54 71.00 43.38 69.45 71.15
T cells(CD3+CD8+) [% of CD3+] 28.36 24.51 29.06 56.73 30.59 28.85
IFN-γ + T cells [% of CD3 + ] 5.74 60.77 0.13 59.62 0.02 3.85
IFN-γ+ T cells [/μl] 131.00 98.30 3.18 0.85 0.49 0.43
IFN-γ + T cells [x10 4 ] 7.86 9.83 0.19 0.08 0.03 0.00
IFN-γ− T cells [% of CD3+] 94.26 39.23 99.87 40.38 99.98 96.15
IFN-γ− T cells [/μl] 2150.00 63.50 2450.00 0.57 2466.67 10.70
IFN-γ− T cells [x104] 129 6.35 147 0.06 148 0.11
IFN-γ+ T cells(CD3+) [% of CD4] 3.83 43.77 0.07 14.53 0.02 1.92
IFN-γ + T cells (CD3+CD4+) [% of CD4] 4.99 56.07 0.06 32.76 0.03 2.70
IFN-γ+ T cells(CD3+CD4+) [/μl] 81.83 68.50 1.04 0.20 0.52 0.21
IFN-γ + T cells abs (CD3+CD4+) [x10 4 ] 4.91 6.85 0.06 0.02 0.03 0.00
IFN-γ+ T cells(CD3+) [% of CD8] 1.80 16.45 0.06 45.09 0.00 1.92
IFN-γ + T cells (CD3+CD8+) [% of CD8] 5.75 64.53 0.21 78.34 0.00 6.67
IFN-γ+ T cells(CD3+CD8+) [/μl] 37.17 25.60 1.50 0.63 0.00 0.21
IFN-γ + T cells abs (CD3+CD8+) [x10 4 ] 2.23 2.56 0.09 0.06 0.00 0.00
C 3. Validation run
Positive control CMVpp65 pp Negative control
OF TCF OF OF TCF OF
volume [ml] 0.6 1.0 0.6 1.0 0.6 1.0
viability [%] 98.57 69.24 98.87 45.75 98.63 26.57
WBCs(CD45+) [x106/ml] 4.23 0.02 3.33 0.01 3.66 0.00
WBCsabs [x106] 2.54 0.02 2.00 0.01 2.20 0.00
T cells(CD3+CD56-) [% of WBCs] 98.13 94.39 99.99 95.45 95.87 73.00
T cells [/μl] 4150.00 21.20 3333.33 11.90 3516.67 0.00
T cellsabs [x106] 2.49 0.02 2.00 0.01 2.11 0.00
T cells(CD3+CD4+) [% of CD3+] 72.20 66.59 72.07 88.51 68.63 84.38
T cells(CD3+CD8+) [% of CD3+] 27.80 33.41 27.93 11.49 31.37 15.62
IFN-γ + T cells [% of CD3 + ] 2.73 70.70 1.49 76.41 0.00 1.56
IFN-γ+ T cells [/μl] 113.33 15.00 49.50 9.06 0.00 0.00
IFN-γ + T cells [x10 4 ] 6.80 1.50 2.97 0.91 0.00 0.00
IFN-γ− T cells [% of CD3+] 97.27 29.30 98.51 23.59 100.00 98.44
IFN-γ− T cells [/μl] 4033.33 6.22 3283.33 2.80 3516.67 0.00
IFN-γ− T cells [x104] 242 0.62 197 0.28 211 0.00
IFN-γ+ T cells(CD3+) [% of CD4] 1.55 45.27 1.35 68.14 0.00 0.76
IFN-γ + T cells (CD3+CD4+) [% of CD4] 2.17 67.26 1.83 76.52 0.00 1.04
IFN-γ+ T cells(CD3+CD4+) [/μl] 65.00 9.50 43.83 8.03 0.00 0.00
IFN-γ + T cells abs (CD3+CD4+) [x10 4 ] 3.90 0.95 2.63 0.80 0.00 0.00
IFN-γ+ T cells(CD3+) [% of CD8] 1.19 26.20 0.18 8.80 0.00 0.52
IFN-γ + T cells (CD3+CD8+) [% of CD8] 4.33 81.11 0.72 78.15 0.00 3.33
IFN-γ+ T cells(CD3+CD8+) [/μl] 50.00 5.75 6.68 1.06 0.00 0.00
IFN-γ + T cells abs (CD3+CD8+) [x10 4 ] 3.00 0.58 0.40 0.11 0.00 0.00

The small-scale MiniMACS CSA was performed as a control for the large-scale CliniMACS CCS process from the same leukapheresis. Detailed information for all three MiniMACS CSA processes (A-B) including viability [%], viable cell number [x106/ml; x106; /μl] and specific cell frequencies [%] for all CSA fractions was determined. The results for the representative analysis of the cells from the original fraction (OF) and T-cell fraction (TCF) are shown. As a positive control, cells were stimulated with SEB, while cells cultured in medium alone served as the negative control; CMVpp65pp = CMVpp65 peptide pool. Bold data reflected the results obtained for the antigen-specific IFN-γ-positive T cells.