Genetically characterized positive control cell lines derived from residual clinical blood samples.
BACKGROUND
Positive control materials for clinical diagnostic molecular genetic testing are in critically short supply. High-quality DNA that closely resembles DNA isolated from patient specimens can be obtained from Epstein-Barr virus (EBV)-transformed peripheral blood lymphocyte cell lines. Here we report the development of a process to (a) recover residual blood samples with clinically important mutations detected during routine medical care, (b) select samples likely to provide viable lymphocytes for EBV transformation, (c) establish stable cell lines and confirm the reported mutation(s), and (d) validate the cell lines for use as positive controls in clinical molecular genetic testing applications.


METHODS
A network of 32 genetic testing laboratories was established to obtain anonymous, residual clinical samples for transformation and to validate resulting cell lines for use as positive controls. Three panel meetings with experts in molecular genetic testing were held to evaluate results and formulate a process that could function in the context of current common practices in molecular diagnostic testing.


RESULTS
Thirteen laboratories submitted a total of 113 residual clinical blood samples with mutations for 14 genetic disorders. Forty-one EBV-transformed cell lines were established. Thirty-five individual point and deletion mutations were shown to be stable after 20 population doublings in culture. Thirty-three cell lines were characterized for specific mutations and validated for use as positive controls in clinical diagnostic applications.


CONCLUSIONS
A process for producing and validating positive control cell lines from residual clinical blood samples has been developed. Sustainable implementation of the process could help alleviate the current shortage of positive control materials.
Bernacki
Susan H
SH
Department of Pathology, Duke University Medical Center, Durham, NC, USA.
Beck
Jeanne C
JC
Stankovic
Ana K
AK
Williams
Laurina O
LO
Amos
Jean
J
Snow-Bailey
Karen
K
Farkas
Daniel H
DH
Friez
Michael J
MJ
Hantash
Feras M
FM
Matteson
Karla J
KJ
Monaghan
Kristin G
KG
Muralidharan
Kasinathan
K
Pratt
Victoria M
VM
Prior
Thomas W
TW
Richie
Kristy L
KL
Levin
Barbara C
BC
Rohlfs
Elizabeth M
EM
Schaefer
Frederick V
FV
Shrimpton
Antony E
AE
Spector
Elaine B
EB
Stolle
Catherine A
CA
Strom
Charles M
CM
Thibodeau
Stephen N
SN
Cole
Eugene C
EC
Goodman
Barbara K
BK
Stenzel
Timothy T
TT
D001800:Blood Specimen Collection; D002461:Cell Line, Transformed; D030342:Genetic Diseases, Inborn / Q000175:diagnosis; D005820:Genetic Testing / Q000379:methods*; D004854:Herpesvirus 4, Human; D006801:Humans; D007753:Laboratories; D008214:Lymphocytes / Q000166:cytology*; D008967:Molecular Biology; D009154:Mutation; D017354:Point Mutation; D017384:Sequence Deletion