[Experimental study on the induction of HLA antibodies by human B lymphocyte cells in vitro].
OBJECTIVE
To explore the optimal in vitro culture condition for producing antibodies to HLA class I molecules by human B lymphocyte cells.


METHODS
By applying EBV-transformed B lymphoblast cell line (BLCL) from a sensitized kidney transplanted subject as a model, the capacity of Ig and specific HLA class I antibody production of BLCL cells was investigated by ELISA and ELISpot, under three different culture systems, RPMI1640+OPTI mAb, RPMI1640+0.1 mmol/L non-essential amino acid+1 mmol/L sodium pyruvate+40 mg/L apo-transferrin,and serum free medium (SFM) for hybridoma culture, and three different culture conditions, culture medium only, culture medium supplemented with IL-4, IL-10 and alpha-CD40, and culture medium supplemented with IL-4, IL-10 and CD40L.


RESULTS
When BLCL cells were cultured in Hybridoma SFM, the IgG and IgM concentrations in culture supernatants were 25.2-28.1 mg/L and 7.14-7.95 mg/L, respectively, which were higher than that of the other two culture systems (P<0.05). Moreover, when Hybridoma SFM was supplemented with IL-4, IL-10 and alpha-CD40 or CD40L, the frequentcies of HLA-I antibody-secreting BLCL cells were 3.06/10(4) BLCL cells and 3.55/10(4) BLCL cells,which were much higher than that of SFM only group (1.77/10(4) BLCL cells).


CONCLUSIONS
BLCL cells can produce both higher amount of Ig and specific HLA-I antibodies under Hybridoma SFM supplimented with IL-4, IL-10 and alpha-CD40 or CD40L.
Liao
Ai-hua
AH
Center of Reproductive Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China. aihualiao@hotmail.com
Vera
Rebmann
R
Hans
Grosse-Wilde
GW
Wang
Yan
Y
Zhang
Ling
L
D000906:Antibodies / Q000276:immunology / Q000378:metabolism*; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D019013:CD40 Antigens / Q000494:pharmacology; D023201:CD40 Ligand / Q000494:pharmacology; D002460:Cell Line; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006680:HLA Antigens / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D007075:Immunoglobulin M / Q000378:metabolism; D016753:Interleukin-10 / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology
D000906:Antibodies; D019013:CD40 Antigens; D016895:Culture Media, Serum-Free; D006680:HLA Antigens; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D016753:Interleukin-10; D023201:CD40 Ligand; D015847:Interleukin-4