Determining cross-reactivity of the clones included in the Human LEGENDScreen™ kit with each of the non-human primate species is only one factor important when preparing for an assay. Identifying any differences or changes in the patterns of expression for each marker on different hematologic cell subsets is equally important for planning an assay or interpreting the results. Since all of the antibodies offered in LEGENDScreen™ are conjugated to phycoerythrin (PE), a panel of backbone markers listed in Table 1. can be employed on complementary fluorophores to PE to phenotype the different subsets. These are clones with known cross-reactivity and expression patterns to best phenotype granulocytes, NK cells, B-cells , T-cells and monocytes in all the NHP species tested. Analysis was done using Cytobank software.

Table 1
CD3 SP34.2 Brilliant Violet 421™
CD20 2H7 Brilliant Violet 605™
CD66 TET2 APC-Vio770®
CD11b ICRF44 PerCP/Cy5.5
CD7 M-T701 APC
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Cells were first gated on CD66abce vs. side scatter to identify granulocytes. From the negative gate, analysis of CD20 versus CD3 yields gates for B-cells and T-cells respectively. From that negative gate, CD7 versus CD11b yields NK cells and monocytes/dendritic cells respectively. From each cell subset, cross-reactivity is determined by PE histograms for the marker in the LEGENDScreen™ kit. As an example, CD2 reactivity for each cell subset is shown here. Each of the NHP species was run in duplicate and compared against human expression.

 

*CD11b+ DCs were not reported independently of the monocytes in this analysis.

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