Secondary LC cluster formation and CD1a expression by cells cultured in 2° cultures in the presence of a panel of mAbs to adhesion molecules.

Purified CD34⁺ cells were cultured in the presence of GM-CSF plus TNFα, SCF, FL, and TGF-β1 in serum-free medium (see “Materials and methods”). Day 8–generated cells were replated in 2° cultures at a high cell density (see “Materials and methods”). After a 2° culture period of 24 hours, cells were harvested and analyzed by phase-contrast microscopy and FACS. The effect of mAb addition on cell cluster formation and CD1a expression density of cultured cells is analyzed (see “Materials and methods”). (A) Bars represent cell cluster formation in one representative experiment (n = 5) quantified using the following scoring system (percentage of cells in aggregates): 0, less than 10%; 1, 10% to 50%; 2, 50% to 75%; 3, 75% to 90%; 4, more than 90%. (B) CD1a mean fluorescence intensity (MFI) of cells after culture in the presence or absence of mAbs.