IMMUNOFLUORESCENCE STUDY OF THE CYTOSKELETON OF A HUMAN GLIOMA CELL LINE BT_(325) / 解剖学报

ABSTRACT

The cytoskeleton of BT_(325), a human glioma cell line, was studied by immunofluorescence microscopy. The effects of different fixatives and buffers on microtubules, microfilaments and intermediate filaments were also compared. It was observed that besides microtubules and microfilaments all cells expressed vimentin. However, only a small fraction of the cells were GFAP positive. We conclude that vimentin is the main component of the intermediate filaments in BT_(325) cells. It was also observed that methanol fixation and formaldehyde fixation followed by acetone or Triton X-100 treatment gave rise to satisfactory results for microtubule immune-staining, and methanol or acidic alcohol fixation resulted in bright staining of intermediate filaments. Formaldehyde fixation also resulted in excellent staining for mierotubnles, but weaker staining for intermediate filaments.If the cells were immune-stained after treatment with Triton X-100, the composition of the buffers had profound effects on microtubules and intermediate filaments, but less effects on microfilaments.It was also observed that if Triton-treated ceils were fixed with formaldehyde before immunostaining with monoclonal antibodies to vimentin, the staining was very weak. However, if the ceils were immunostained first and then fixed with for-maldehyde the staing was quite bright. We conclude that for-maldehyd e fixation may "mask" or destroy certain epitopes on vimentin molecules.