We have obtained a cDNA fragment to human glial fibrillary acidic protein (GFAP) by immunoscreening a lambda gt11 human brain cDNA library with antibody to bovine GFAP. The highly homologous nucleotide sequence of this clone with that of the mouse GFAP enabled the identification of this cDNA as one encoding GFAP. As this cDNA hybridized with a single major RNA species in Northern blots of RNA from human and mouse brain tissues and gave one or two bands in Southern blots of human genomic DNA, it was considered to be specific for GFAP. Using this cDNA as a probe we investigated the levels of GFAP expression in ten human glioma cell lines. A 3.5-kb GFAP mRNA was detected in five of the ten glioma cell lines, one of which was U-251 MG cell line and the other four were clones derived from the same tumor (CL1, 2, 3, and 4). There was a difference in the amount of GFAP mRNA among U-251 MG and the four clonal cell lines. Quantitative evaluation of this difference by RNA dot blot analysis revealed that the amount of GFAP mRNA expressed in CL3 was about 1/5 and in CL4 about 1/10 the amount expressed in U-251 MG, CL1, and CL2. Semiquantitative Western blot analysis showed that GFAP levels corresponded to the GFAP mRNA levels in these cell lines. By Southern blot analysis of genomic DNA the GFAP gene was similarly detected in all of these cell lines regardless of the level of GFAP expression.(ABSTRACT TRUNCATED AT 250 WORDS)