Loss of heterozygosity for chromosome 13q including the RB locus is a common genetic alteration in small-cell lung carcinoma (SCLC) as well as in retinoblastoma. We examined the RB cDNA sequences of exon 13 to 18 and exon 19 to 23 in 9 SCLC cell lines to detect mutations which cause inactivation of the remaining allele of the RB gene. Internal deletions of RB cDNA were observed in 3 of the 9 SCLC cell lines. In the Lu-24 cell line, a 114 base pairs (bp) deletion corresponding to exon 22 was due to abnormal splicing, which probably resulted from a two-base mutation within genomic exon 22, and aberrant 105 kilodaltons RB protein was detected by immunoprecipitation analysis. A base pair deletion within exon 20 in the Lu-135 cell line and a 1 bp deletion within exon 23 in the Lu-141 cell line were due to the deletions of the corresponding genomic DNA, and each deletion resulted in formation of a premature termination codon. These results indicate that both alleles of the RB genes are inactivated in SCLC by several different mechanisms, including small deletion, mutation and chromosomal loss.