Figure 2

Western analysis with SMI-31, RT97, anti-lamin B, SMI-32 on SH-SY5Y, and SMI-31, RT97, anti-lamin B, anti-GFAP on F98 cell extracts subjected to SDS-PAGE. On SH-SY5Y, the SMI-31 and SMI-32 lanes show strong labeling of bands corresponding to the MW of NF-H, and NF-M. Additional bands, not seen in the SMI-32, are present in the SMI-31 and RT97 lanes between and above the NF-H and NF-M, and probably represent phosphorylated forms of NF-M and NF-H. SMI-31, RT-97 and anti-lamin B also revealed a band or closely spaced bands at approximately 70 kD which corresponds approximately to the MW of lamin B. Western analysis with anti-lamin A/C revealed no bands in either SH-SY5Y or F98 extracts (not shown). On F98 cells, SMI-31 labeled several well defined bands, including those of MW corresponding to GFAP and lamin B. The RT97 mAB also strongly labeled several F98 bands, including bands corresponding to GFAP and lamin B. The anti-lamin B mAB strongly labeled bands at the appropriate MW (~70 kD) and also a cluster of bands in the range of the heavy NF MW (~207 kD). The anti-GFAP pAB labeled a band at the appropriate MW (~52 kD) and also band higher of MW appropriate for multimers of GFAP (~100, 150, 200 kD). Labeled bands lower than 65 kD may represent degradation products produced during cell extraction. Negative controls (control blots stained with appropriate normal serum) showed no labeling (data not shown).