Mutation analysis of patient 7. (A) Heterozygous germline mutation in exon 3 of CD95 causing a 68Pro → 68Leu change. Genomic DNA of PBMC was subjected to PCR using primers flanking exon 3 of CD95, amplificates were subjected to SSCP analysis, abnormal migrating bands were cut-out from the gel, reamplified, and sequenced. T-ALL blasts and mature T lymphocytes were immunomagnetically purified, exon 3 of CD95 amplified by PCR and sequenced. Nucleotide sequence is compared to wild type. Protein is numbered in respect to the start of the mature protein (according to Itoh et al46). (B) CD95 mRNA is strongly expressed compared with cryopreserved blasts from a different patient and thawed Jurkat cells. mRNA was determined by RT-PCR and compared with GAPDH.