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: Sequencing can be performed by either Sanger Sequencing or Next-Generation Sequencing.
Sanger Sequencing: Sequencing of CFTR is carried out by amplification of all 27 exons and intron/exon boundaries followed by bi-directional Sanger sequencing. The sensitivity of full gene sequencing is estimated to be approximately 99% for single nucleotide substitutions and small insertions/deletions. All nucleotide changes are analyzed within the context of current databases and literature to predict pathogenicity.
NGS: Next generation sequencing will analyze the exons or coding regions of CFTR using Illumina NextSeq 500 technology. Samples are prepared using hybridization probes to enrich exonic regions. Promoter, intronic, etc. regions are not assessed on our assay, but may contain variants that impact gene function.
Component 2: Large deletions and duplications can be detected by either MLPA or Exon-Centric Microrray.
MLPA: Reflex multiplex ligation-dependent probe amplification (MLPA) will be used to detect large CFTR coding region deletions /duplications when indicated.
Exon-Centric Microrray: Alternatively, a customized CytoSure ‘Exon-centric’ array (Oxford Gene Technology) may be used for exon-level deletion and duplication analysis. The targeted array has enhanced probes targeted to the exonic regions of CFTR. The arrays are run using Agilent SureScan technology. Any statistically-significant genetic changes are compared against all known databases and relevant literature. Test reporting follows the ACMG Standards & Guidelines for Clinical Genetics Laboratories, Ultra-Rare Disorders Guidelines, and Interpretation of Sequence Variants Guidelines.
Specimen Requirements:
Blood: Lavender tube (EDTA) or yellow (ACD)
- Adult: 6.0 mL
- Child: 6.0 mL
- Infant: 2.0 mL
Prenatal:
- Direct Amniotic Fluid (10-20mL)
- Direct CVS
- Cultured Amnio or CVS (2-T25 flasks)
DNA: 10µg at a minimum of 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 CVS or Amniotic Fluid Specimens:
- Maternal cell rule-out testing will be performed on all prenatal specimens received. Please provide maternal blood in addition to the fetal specimen. Additional charges apply for the maternal cell rule-out test.
- All genetic testing performed on Direct CVS or Amniotic Fluid specimens will be confirmed on cell cultures prepared by Knight Diagnostic Laboratories. Cell cultures will be prepared from the specimen received. Additional charges apply for confirmatory testing.
For routine testing of blood, saliva and buccal swabs, 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 outside of 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.