Figure 4.

Lymphocytes after the two-step fusion procedure and before separation by velocity sedimentation. (a) Cells were cytocentrifuged on a glass slide and stained by May-Grünwald Giemsa. Bi- and trinucleated cells (black arrows) were readily observed (800×). (b) Cell fusion products, in vivo, were transferred into an Iwaki quartz base dish and observed by inverted fluorescence microscopy using a dual filter (485/515 nm and 578/610 nm) and by phase contrast microscopy (500×). One fusion product, at the bottom, is entirely green (two CMTMR-labeled cells fused) while the three fusion products (white arrow) in the centre-left are yellow (one CMTMR-labeled cell fused with a CMFDA-labeled cell). The mixing of the two fluorescent dyes takes some time and it is mainly dependent on temperature and the viscosity of cell cytoplasm. Yellow fluorescent is evident only when the two fluorescent dyes, CMTMR and CMFDA, are completely mixed.