[Experimental study on the induction of HLA antibodies by human B lymphocyte cells in vitro]

To explore the optimal in vitro culture condition for producing antibodies to HLA class I molecules by human B lymphocyte cells.

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.

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).

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.