Background:
Cystic Fibrosis (CF) is the most common autosomal recessive genetic disease in the Caucasian population and 1 appears in approximately one in every 2,500 newborns. In the Caucasian and Ashkenazi Jewish populations, about one in every 25 individuals carries one copy of a mutation in the CFTR gene at 7q31.2.
Reasons for Referral:
- Confirmation of clinical diagnosis
- Carrier identification in persons with a positive or negative family history
- Sperm & egg donors
- Abnormal fetal ultrasound
- Preconception and carrier screening
Methodology:
This test has two components:
Component 1: Genomic DNA is analyzed using next-generation sequencing (NGS) on the Illumina NextSeq 2000 platform, with target enrichment performed using hybridization-based probes to capture exonic (coding) regions of the gene(s). Single nucleotide variants (SNVs) and small insertions or deletions (INDELs) are identified using the Illumina DRAGEN Enrichment Workflow, executed onboard the NextSeq2000. This pipeline combines software and hardware acceleration to generate high-confidence germline haplotype calls. Clinical and analytical validation of DRAGEN was performed in our laboratory. Based on validation study results, for SNVs, this assay achieves >96% analytical sensitivity and >99% positive predictive value (PPV). For INDELs 87% and the PPV is >97%. INDELs >50 bp may be detected but the sensitivity for these is reduced.
This test is not designed to detect polynucleotide repeats, low-level mosaicism, structural rearrangements or balanced alterations (e.g. inversions, gene conversion events, translocations, etc.) or variants in difficult regions. Additionally, variants located in regions of insufficient coverage, including introns and promoter regions; pseudogenes; where the reference genome is inaccurate or contains gaps and insertions; and of high GC content may not be detected. This test does not provide complete coverage of all exons and noncoding regions may have limited information and ability to interpret. Variants in introns that are greater than 10 bp from the intron-exon junction may be analyzed. Please contact the laboratory if interrogation of intronic sequence greater than 10 bp from the intron-exon boundary is desired.
Component 2: Multiplex ligation-dependent probe amplification (MLPA) to detect large CFTR coding region deletions /duplications is performed. The frequency of large deletions in the CFTR gene is estimated to be 1-3% (Schneider M. et al).
Specimen Requirements:
Blood: Lavender tube (EDTA) or yellow (ACD)
- Adult: 6.0 mL
- Child: 6.0 mL
- Infant: 2.0 mL
Saliva: 2 ORAgene Saliva Kits (OGR-500) used according to manufacturer instructions. Please contact KDL Client Services for a Saliva Collection Kit for patients that cannot provide a blood sample.
Assisted Saliva: 4 ORAgene Assisted Saliva Kits (OGR-575) used according to manufacturer instructions. Please contact KDL Client Services for an Assisted Saliva Collection Kit for patients that cannot provide a blood sample.
Buccal Cells: 4 CytoSoft™ Cytology Brush (Medical Packaging CYB-1) used according to manufacturer instructions. Please contact KDL Client Services for a Buccal Collection Kit for patients that cannot provide a blood sample.
Skin Fibroblast: Punch Biopsy (cell cultures will be prepared at KDL and used for testing), or 2 T-25 confluent flasks.
DNA: 5-10µg at a minimum of 60-100ng/µL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS).
Prenatal:
- Direct Amniotic Fluid (10-20mL)
- Direct CVS
- Direct POC
- Cultured Amniocytes (2-T25 flasks)
- Cultured CVS (2-T25 flasks)
- Cultured fetal tissue: Product of Conception (2 T25 flasks)
- Cord Blood (1-2mL)
DNA: 5-10µg at a minimum of 60-100ng/µL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS).
Notice Regarding Molecular Genetic Testing on Prenatal Specimens:Maternal cell rule-out testing will be performed on all prenatal specimens received. Please provide maternal blood (or saliva) in addition to the fetal specimen. Additional charges apply for the maternal cell rule-out test.
For routine testing of blood and saliva (or DNA extracted from them), KDL does NOT accept samples from patients within two (2) weeks of a packed cell/platelet transfusion or within four (4) weeks of a whole blood transfusion. For extraordinary circumstances, where testing must be performed within the above windows, please contact our lab.
A REQUISITION FORM MUST ACCOMPANY ALL SAMPLES. Please include detailed clinical information, including ethnicity, clinical history, and family history.
Test Performed (Days):
Weekly
Turn Around Time:
14 – 21 Days
Shipment Sensitivity Requirements:
- Keep specimen cold during transit, but do not ship on dry ice.
- Please use the cold pack provided in the KDL shipping kit.
- Ship the specimen overnight express, using the FedEx priority overnight label provided.
References:
Additional Info:
Prior to any genetic testing we recommend genetic counseling. To receive forms and information about prenatal diagnostic testing, please contact Client Services at (855) 535-1522.