Patients. Between 1994 and 2001, a total of 171 patients were referred to the Genetic Unit of the 1^st De­partment of Pediatrics at Athens University, Athens, Greece, mainly from pediatricians, neurologists and genet­icists for molecular investigation of PWS (114) and AS (57) syndromes. Most of the patients were clinically eval­uated, and classified according to specific criteria, as having typical or atypical clinical presentation [1,5]. Pa­tients with hypotonia, characteristic faces, hypogonadism and cytogenetic and/or molecular abnormality, who were younger than 3 years or hypotonia in the neonatal period, obesity in early childhood, hypogonadism, short stature, learning difficulties and characteristic face after the age of 3 years, were considered as typical PWS. All the re­maining cases were classified as non typical [1]. As typi­cal AS were considered the patients with severe develop­mental delay, lack of speech, ataxic gait and improper laughter, while the ones with some of these characteristics were considered as non typical. Family history was avail­able in most cases. Samples from the patient and both parents were analyzed. Twelve patients referred for PWS and six for AS were not examined, since only blood sam­ples and short clinical reports were sent from other centers.

Prader Willi syndrome referrals comprised 79 males and 35 females, ranging in age from 7 days to 16 years (mean age 5.5 years). Angelman syndrome referrals were 22 males and 35 females, and their age ranged from 13 months to 21 years (mean age 7.6 years).

Cytogenetic and Molecular Cytogenetic Analysis. High-resolution cytogenetic analysis on prometaphase chromosomal spreads was performed in all patients for whom fresh blood samples were available (63/171) (Table 1). Fluorescent in situ hybridization (FISH) analysis using the SNRPN probe on the 15q11.13 region (Appligene Oncor; Heidelberg, Germany) was applied selectively in 34/114 cases with uninformative molecular investigation or karyotype abnormality (Table 1).

Molecular Investigation. Genomic DNA was ex­tracted from peripheral blood lymphocytes using the salt­ing out method. Dinucleotide repeat polymorphism (DNRP) analysis was performed with the use of six primer sets mapped within the 15q11-13 regions: D15S10, D15S11, D15S113, GABRb3, D15S543, and D15S97. To test UPD, two loci mapped outside the region towards the telomere, D15S87 and CYP19 were analyzed [9-11]. Polymerase chain reaction (PCR) conditions were as fol­lows: for D15S10 94°C 1 min., 52°C 2 min., 72°C 2 min. (30 cycles); for D15S11, D15S113 and GABRb3, 93°C 1 min., 58°C 2 min., 72°C 2 min. (26 cycles); for D15S543, 94°C 1 min., 56°C 1 min., 72°C 1 min. (30 cycles); D15S97, 94°C 1 min., 60°C 2 min., 72°C 1 min. (30 cycles); for D15S87, 94°C 1 min., 55°C 2 min., 72°C 2 min. (27 cycles); and for CYP19, 94°C 1 min., 54°C 1 min., 72°C 1 min. (30 cycles). Electrophoresis was per­formed on a 6% polyacrylamide/7M urea gel, and diagno­sis was confirmed by the detection of absent paternal al­leles for PWS and absent maternal alleles for AS.