Provision of Epstein-Barr virus-transformed B-cell lines in a routine tissue typing laboratory: practicalities and applications.
A bank of Epstein-Barr virus (EBV)-transformed B-lymphoblastoid cell lines (BCLs) has been established in this laboratory over the last 7 years. Novel, rare and unusual HLA phenotypes were highlighted during routine clinical testing and during typing of volunteer haematopoietic stem cell donors for the Welsh Bone Marrow Donor Registry. The BCLs are routinely used as laboratory reagents, as a source of DNA for reference purposes and for research work. Consenting donors are tested for hepatitis B surface antigens (HBsAg), human immunodeficiency virus (HIV) and hepatitis C virus (HCV) before their B-lymphocytes are transformed. Strict bio-identity checks are performed before and after transformation and after each BCL expansion. Each BCL is tested regularly for mycoplasma contamination. A total of 230 blood samples were transformed. One hundred and fifty-nine sterile samples produced 157 BCLs (98.7% success), while 71 non-sterile samples produced 50 BCLs (70.4% success), giving an overall success rate of 90.0%. Fifteen of the transformation failures have since been repeated successfully. Factors contributing to the high success rate and reasons for the 23 failed transformations are discussed. The successful development and use of pools of BCLs for HLA antibody screening by flow cytometry are described. Whilst certain training and health and safety issues require close attention, it is clear that EBV transformation of B lymphocytes and/or the use of BCLs is feasible in a routine tissue typing laboratory and that BCLs are a valuable resource. Careful adherence to the methods and procedures detailed here should virtually guarantee successful transformation (98.7%) from good quality, sterile whole blood samples.
Bass
H
H
Welsh Transplantation and Immunogenetics Laboratory, Welsh Blood Service, Pontyclun, UK. helen.bass@wbs.wales.nhs.uk
Walters
R
R
Darke
C
C
D001402:B-Lymphocytes / Q000276:immunology* / Q000821:virology; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral; D005434:Flow Cytometry; D006680:HLA Antigens / Q000235:genetics / Q000276:immunology; D004854:Herpesvirus 4, Human / Q000235:genetics* / Q000276:immunology; D006650:Histocompatibility Testing / Q000379:methods*; D006801:Humans; D011786:Quality Control
D006680:HLA Antigens