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Thus, it appears that SMI-31 recognizes a phosphoepitope that occurs on proteins in addition to NF-H and NF-M, as previously reported (see the Discussion for other reports of SMI-31 staining glial cells in situ ). RT-97 also labeled nuclear and cytoplasmic structures in F98 cells (Figure 1E).
Because of the association of phosphorylated lamins with DNA replication and the cell cycle [ 8 ] , and the sharing of phosphoepitopes among lamins, NFs, and GFAP, it seems most probable that the nuclear epitopes revealed in neuroblastoma and glioma cells by SMI-31 and RT-97 are contained on lamin B.
We asked the following questions: 1) are the immuno-labeled structures within the nucleus or just closely associated; 2) is the phosphoepitope labeled by SMI-31 and RT-97 mABs specific to NFs or can it be identified on other IFs in other cell types; and 3) is there a relationship between the cell cycle as determined by DNA synthesis and the amount of nuclear labeling by SMI-31 and RT-97?
Comparison among antibodies that are specific for phosphorylated IFs (SMI-31, RT-97) and those not (SMI-32, anti-GFAP) suggests that the nuclear structures are a phosphorylated IF.
Multiple bands also appear in both the SMI-31 and RT-97 lanes, although for the most part the antibodies do not appear to label the same bands.