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Table 3 Antibody tritation

From: Validation of analytical methods in compliance with good manufacturing practice: a practical approach

Markers Antibody concentration (μl)
  5 10 20 40 80
CD45 FITC 2.0 2.0 2.0 1.9 1.9
CD34 FITC 1.8 1.9 2.0 1.9 1.9
CD14 FITC 1.9 1.9 1.9 1.9 1.9
HLADR PE 0.8 0.7 0.7 0.8 0.9
CD73 PE 1.8 1.9 2.0 2.0 2.1
CD83 FITC 1.7 1.8 1.8 1.9 2.0
CD40 PE 1.5 1.7 1.6 1.7 1.8
CD80 FITC 1.9 1.9 1.9 1.9 2.0
CD86 PE 1.1 1.1 1.1 1.2 1.5
HLADR PercPCy5.5 1.2 1.2 1.3 1.2 1.3
CD45RA FITC/CD45RO PE/CD3 PercP/CD8 APC 1.7 1.9 1.5 1.5 1.7
CD45RA FITC/CD45RO PE/CD3 PercP/CD8 APC 2.6 3.0 2.5 2.0 2.2
CD45RA FITC/CD45RO PE/CD3 PercP/CD8 APC 2.5 2.5 2.5 1.9 2.1
CD45RA FITC/CD45RO PE/CD3 PercP/CD8 APC 2.4 3.3 2.6 2.5 1.7
CD45RA FITC/CD45RO PE/CD3 PercP/CD4 APC 1.8 1.7 1.4 1.3 1.4
CD45RA FITC/CD45RO PE/CD3 PercP/CD4 APC 2.2 2.1 2.2 2.2 2.1
CD45RA FITC/CD45RO PE/CD3 PercP/CD4 APC 2.5 2.4 2.4 2.2 2.1
CD45RA FITC/CD45RO PE/CD3 PercP/CD4 APC 5.1 5.0 4.5 4.6 4.5
CD3 FITC/CD8 PE/CD4 APC/CD45 PercP 2.2 2.0 2.0 2.0 1.9
CD3 FITC/CD8 PE/CD4 APC/CD45 PercP 2.6 1.8 1.8 1.2 1.2
CD3 FITC/CD8 PE/CD4 APC/CD45 PercP 4.3 4.2 4.0 4.0 4.0
CD3 FITC/CD8 PE/CD4 APC/CD45 PercP 4.0 3.9 3.8 3.7 3.6
CD3 FITC/CD56-16 PE/CD19 APC/CD45 PercP 2.1 2.4 2.4 2.3 2.1
CD3 FITC/CD56-16 PE/CD19 APC/CD45 PercP 0.9 1.2 1.6 1.2 1.1
CD3 FITC/CD56-16 PE/CD19 APC/CD45 PercP 2.8 1.8 3.4 2.4 1.9
CD3 FITC/CD56-16 PE/CD19 APC/CD45 PercP 3.1 3.3 3.7 3.7 3.7
Markers Antibody concentration (μl)
  2 5 10 20 40
CD105 APC 1.8 1.8 1.8 1.8 1.8
CD90 FITC 1.8 1.8 1.9 1.9 1.8
Markers Antibody concentration (μl)
  1 2 5 10 20
CD11c APC 1.6 1.4 1.3 1.4 1.4
CD19 APC 2.3 2.8 2.9 2.7 2.4
  1. The immunophenotype validation required a first step which is the tritration of each antibody performed by using scalar antibody dilution. The best antibody concentration was that with a higher resolution index. The lowest antibody concentration was chosen when there was an equal resolution index.