Can the Diagnosis of NF1 Be Excluded Clinically?
Can the Diagnosis of NF1 Be Excluded Clinically?
Affected individuals were ascertained through the Manchester nationally commissioned service for complex NF1 and examined (by DGRE, SMH or OQ) in its specialist clinics (based in Manchester). Where indicated, MRI scans of the neuroaxis and/or whole body were carried out, and interpreted along with the clinical phenotypes in the context of a multidisciplinary team discussion. Permission for inclusion of clinical and imaging details was sought from the patients reported here.
Molecular analyses were carried out in the clinical pathology accredited Manchester Regional Genetics Laboratory and in the Genetics Research and Development laboratory in Cardiff. RNA and genomic DNA were prepared from peripheral blood samples. RNA was reverse transcribed to cDNA using standard procedures, and direct sequencing performed to demonstrate splicing abnormalities or mutations within the coding sequence. Mutation status was confirmed in genomic DNA. Multiplex ligation dependent probe amplification (MLPA) for dosage analysis was additionally performed in samples without a clearly pathogenic mutation; for example, where a novel sequence variant with uncertain pathogenicity had been identified.
Methods
Affected individuals were ascertained through the Manchester nationally commissioned service for complex NF1 and examined (by DGRE, SMH or OQ) in its specialist clinics (based in Manchester). Where indicated, MRI scans of the neuroaxis and/or whole body were carried out, and interpreted along with the clinical phenotypes in the context of a multidisciplinary team discussion. Permission for inclusion of clinical and imaging details was sought from the patients reported here.
Molecular analyses were carried out in the clinical pathology accredited Manchester Regional Genetics Laboratory and in the Genetics Research and Development laboratory in Cardiff. RNA and genomic DNA were prepared from peripheral blood samples. RNA was reverse transcribed to cDNA using standard procedures, and direct sequencing performed to demonstrate splicing abnormalities or mutations within the coding sequence. Mutation status was confirmed in genomic DNA. Multiplex ligation dependent probe amplification (MLPA) for dosage analysis was additionally performed in samples without a clearly pathogenic mutation; for example, where a novel sequence variant with uncertain pathogenicity had been identified.
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