Summary Specimen Handling and Human DNA Isolation in Molecular Genetics Clinical Lab

Specimen Procurement
 Specimen collection = PB  trisodium EDTA (purple)  Preferred anticoagulant in tubes
 Acid citrate dextrose (yellow)  For Paternity testing
 Frozen specimen
o AF or CVS
 Direct/cultured isolation
 CVS  Arrive in heparin tubes if collected by cytogenetics lab
 AF  Usually cultured before molecular assays are attempted
 Culturing should be completed (>75% confluency)
 All prenatal specimens  Collect maternal blood at the same time so maternal cell
contamination can be assessed
o Not accepted at BCCH  Buccal swab, saliva, FFPE specimen, blood spot cards

 CLSI Storage of Specimens Guidelines
o DNA
 PB
 RT = 24 hrs, 4℃ = 3 days
 CVS/AF (Direct sampling)
 Isolate ASAP/day of arrival/4℃ overnight, extraction next day
 CVS/AF (Cultured)
 Isolate ASAP/within 2 hours of arrival/store at 4℃
 DNA in TE Buffer
 RT = up to 26 weeks; 4℃ = up to 1 year; -20 = up to 7 yrs, -70 > 7 yrs
 Blood is the only specimen that can be kept at RT for extended periods (>1 hr, up to 8
days) without loss/degradation of DNA
 Do not freeze whole blood
o RNA
 PB
 Collection tube should have RNA stabilization additive
 Isolate within 4 hrs of arrive/store isolated PB at -70℃
 CVS/AF (Direct sampling)
 Flash freeze immediately after collection
 Isolate RNA within 4 hrs of arrival/store at -70℃
 CVS/AF (Cultured)
 Isolate RNA within 4 hrs of arrival/store at -70℃
 RNA in ethanol
 Only store in -70℃
o Ultrahigh Molecular Weight DNA (UHMW DNA)
 Min. 150kb to Mb sized fragments; DNA must be clean
 Fresh Blood
 RT (20-25℃) Up to 66 hrs; 6 hrs max. time for higher temp.
 4℃ 5 days max.