Establishment of stably EBV-transformed cell lines from residual clinical blood samples for use in performance evaluation and quality assurance in molecular genetic testing.
Positive control materials for clinical molecular genetic testing applications are currently in critically short supply or non-existent for many genetically based diseases of public health importance. Here we demonstrate that anonymous, residual, clinical blood samples are potential sources of viable lymphocytes for establishing Epstein-Barr virus (EBV)-transformed blood lymphocyte cell lines. We attempted to transform 34 residual blood samples, and analyzed transformation success with respect to sample age, anticoagulant, storage temperature, volume, hemolysis, and patient age and sex. In univariate analysis, sample age was significantly associated with transformation success (P = 0.002). The success rate was 67% (6 of 9) for samples 1 to 7 days old, 38% (3 of 8) for samples 8 to 14 days old and 0% for samples 15 to 21 (0 of 11) days old. When we controlled for sample age in multivariate logistic regression, anticoagulant and storage temperature approached significance (P = 0.070 and 0.087, respectively; samples in acid citrate dextrose (ACD) and refrigerated samples were more likely to transform). Based on these findings, we suggest that samples collected in either ACD or ethylene diamine tetraacetic acid, and up to 14 days old (refrigerated) or 7 days old (stored ambient), are reasonable candidates for EBV transformation. The transformation rate for samples that met these criteria was 63% (10 of 16). Implementation of this process could help alleviate the shortage of positive control materials for clinical molecular genetic testing.
Molecular Diagnostics Laboratory, Duke University Medical Center, Durham, North Carolina, USA.
Susan H
SH
Bernacki
Ana K
AK
Stankovic
Laurina O
LO
Williams
Jeanne C
JC
Beck
James E
JE
Herndon
Karen
K
Snow-Bailey
Thomas W
TW
Prior
Karla J
KJ
Matteson
Linda M
LM
Wasserman
Eugene C
EC
Cole
Timothy T
TT
Stenzel
D000328:Adult; D000375:Aging; D000925:Anticoagulants / Q000494:pharmacology; D001800:Blood Specimen Collection; D018929:Cell Culture Techniques / Q000379:methods*; D002461:Cell Line, Transformed; D005069:Evaluation Studies as Topic; D005260:Female; D005820:Genetic Testing / Q000379:methods / Q000592:standards*; D004854:Herpesvirus 4, Human / Q000502:physiology*; D006801:Humans; D008214:Lymphocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism / Q000821:virology*; D008297:Male; D008875:Middle Aged; D008967:Molecular Biology / Q000379:methods / Q000592:standards*; D011786:Quality Control; D012727:Sex Characteristics; D013696:Temperature; D013997:Time Factors
D000925:Anticoagulants