MITOCHONDRIAL DNA DEFECTS
Mitochondrial DNA (mtDNA) defects are classified into three groups: point mutations, large-scale rearrangements (deletions and duplications), and depletion. mtDNA point mutations are detected by polymerase chain reaction (PCR) amplification of regions of mtDNA followed by restriction fragment length polymorphism (RFLP) analysis. Positive results are confirmed by direct sequencing of the regions of interest. Large-scale rearrangements (deletions and duplications) of mtDNA are detected by Southern blot hybridization technique. In this method, mtDNA is digested with restriction enzyme(s), the fragments are separated by electrophoresis, and are then transferred onto a nylon membrane for hybridization with an appropriately radiolabeled probe. The hybridized mtDNA is visualized by autoradiography.
Depletion of mtDNA is detected by quantitative real-time PCR (q-PCR) assay. In this assay, a segment of mtDNA and a single copy nuclear gene (used as an internal standard for quantification) are simultaneously amplified and measured, using specific primers and fluoregenic probes.
Specimen requirement:
Blood:
We require approximately 6ml of blood in tubes containing ACD citrate solution (yellow top).
Muscle biopsies:
Muscle biopsies from any site are acceptable. The most common sites are thigh or arm. The muscle should be snap frozen immediately in liquid nitrogen and shipped on dry ice. The biopsy should be approximately 0.1 cm in diameter and 0.5 cm in length; approximate weight should be 100-200 mg
Shipping:
Ship specimens via an overnight courier in a styrofoam container with 5 to 10 lbs of dry ice. Shipping time in excess of 24 hrs will result in failure to maintain appropriate temperatures and affects the results of the assay.
The blood should be sent at room temperature to the address below:
The muscle should be sent frozen, on dry ice, by overnight carrier to the address below:
Dr. Ali Naini
Laboratory of Melocular Neurogenetics
Columbia University Medical Center
Department of Pathology
630 West 168th Street, VC15th Floor, Room 208
New York, NY 10032
Phone: 212-305-3947
FAX: 212-305-3986
CPT Codes:
83890, 83892, 83894, 83898, 83912 (per point mutation).
Price:
$225 per point mutation |
