Purification to homogeneity and characterization of human B-cell differentiation factor (BCDF or BSFp-2).

Human B-cell differentiation factor (BCDF) was purified to homogeneity by sequential filtration and chromatography of culture supernatants from TCL-Na1 cells on an AcA34 gel column and then on a Mono P column with fast protein liquid chromatography and reversed-phase HPLC. A 5300-fold enrichment in specific activity of BCDF with about 25% recovery was attained. The homogeneity of purified BCDF was evidenced by the following: (i) the specific activity was 1.7 X 10(7) units/mg of protein, (ii) only two bands, Mr 19,000 and 21,000, were identified by NaDodSO4/PAGE under reduced as well as nonreduced conditions, and (iii) BCDF activity was recovered from the gel after NaDodSO4/PAGE in the fractions corresponding to protein bands of Mr 19,000 or 21,000. Purified BCDF induced Ig secretion in Epstein-Barr virus-transformed cell lines; as little as 3 pM gave 50% of the maximum reaction achieved by 30-80 pM BCDF. Purified BCDF induced Ig production in activated B cells without any effect on cell growth. Purified BCDF did not show any activity of interleukin 1 or 2, B-cell stimulatory factor (BSF)p-1, B-cell growth factor II (BCGF-II), or interferon. Since BCDF was isolated and characterized as described, we propose that the BCDF that induces the final differentiation of B cells into high-rate Ig-secreting cells be designated BSFp-2.

[1]  Kendall A. Smith,et al.  Heterogeneity of human T-cell growth factor(s) due to variable glycosylation. , 1981, Molecular immunology.

[2]  E. Wecker,et al.  Replacement of T-cell function by a T-cell product. , 1972, Nature: New biology.

[3]  T. Hirano,et al.  Human helper T cell factor(s). IV. Demonstration of a human late-acting B cell differentiation factor acting on Staphylococcus aureus Cowan I-stimulated B cells. , 1984, Journal of immunology.

[4]  E. Puré,et al.  Induction of B cell differentiation by T cell factors. I. Stimulation of IgM secretion by products of a T cell hybridoma and a T cell line. , 1981, Journal of immunology.

[5]  P. Marrack,et al.  Role of γ-interferon in antibody-producing responses , 1984, Nature.

[6]  T. Hirano,et al.  Effect of recombinant IL 2 and gamma-IFN on proliferation and differentiation of human B cells. , 1985, Journal of immunology.

[7]  R. Mertelsmann,et al.  Human B cell-inducing factor(s) for production of IgM, IgG and IgA: independence from IL 2. , 1984, Journal of immunology.

[8]  S. Swain,et al.  Culture supernatants of a stimulated T-cell line have helper activity that acts synergistically with interleukin 2 in the response of B cells to antigen. , 1981, Proceedings of the National Academy of Sciences of the United States of America.

[9]  D. Vaux,et al.  Proliferation and differentiation of single hapten-specific B lymphocytes is promoted by T-cell factor(s) distinct from T-cell growth factor. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[10]  T. Kishimoto,et al.  T cell-replacing factor- (TRF) induced IgG secretion in a human B blastoid cell line and demonstration of acceptors for TRF. , 1981, Journal of immunology.

[11]  W. Paul,et al.  Soluble factors involved in B cell differentiation: identification of two distinct T cell-replacing factors (TRF). , 1983, Journal of immunology.

[12]  T. Hirano,et al.  Demonstration of the involvement of interleukin 2 in the differentiation of Staphylococcus aureus Cowan I-stimulated B cells. , 1984, Journal of immunology.

[13]  T. Hirano,et al.  Human helper T cell factor(s). III. Characterization of B cell differentiation factor I (BCDF I). , 1984, Journal of immunology.

[14]  P. Goldsmith.  A highly sensitive enzyme-linked immunosorbent assay for human immunoglobulin E: comparison of microtiter plate and disk methodologies. , 1981, Analytical biochemistry.

[15]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[16]  T. Kishimoto,et al.  Induction of proliferation and Ig production in human B leukemic cells by anti-immunoglobulins and T cell factors. , 1982, Journal of immunology.

[17]  S. Akira,et al.  Effect of B cell differentiation factor (BCDF) on biosynthesis and secretion of immunoglobulin molecules in human B cell lines. , 1985, Journal of immunology.

[18]  P. Ralph,et al.  Clonal differences in response to T cell replacing factor (TRF) for IgM secretion and TRF receptors in a human B lymphoblast cell line , 1983, European journal of immunology.

[19]  T. Kishimoto,et al.  Isolation and characterization of B cell differentiation factor (BCDF) secreted from a human B lymphoblastoid cell line. , 1984, Journal of immunology.

[20]  A. Tominaga,et al.  Antigen-induced T cell-replacing factor (TRF). III. Establishment of T cell hybrid clone continuously producing TRF and functional analysis of released TRF. , 1980, Journal of immunology.

[21]  P. Marrack,et al.  Evidence for two distinct classes of murine B cell growth factors with activities in different functional assays , 1983, The Journal of experimental medicine.

[22]  K. Dobashi,et al.  Purification and physicochemical characterization of murine T cell replacing factor (TRF). , 1985, Journal of immunology.

[23]  P. Marrack,et al.  B cell helper factors. I. Requirement for both interleukin 2 and another 40,000 mol wt factor , 1981, The Journal of experimental medicine.

[24]  W. Paul,et al.  Regulation of B-cell growth and differentiation by soluble factors. , 1983, Annual review of immunology.

[25]  W. Paul,et al.  Identification of a T cell-derived b cell growth factor distinct from interleukin 2 , 1982, The Journal of experimental medicine.

[26]  A. Fauci,et al.  B cell growth factor and T cell growth factor produced by mitogen-stimulated normal human peripheral blood T lymphocytes are distinct molecules. , 1982, Journal of immunology.

[27]  S. Pestka,et al.  A convenient and rapid cytopathic effect inhibition assay for interferon. , 1981, Methods in enzymology.

[28]  M. Schreier,et al.  Mediators from cloned T helper cell lines affect immunoglobulin expression by B cells. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[29]  T. Hirano,et al.  Human helper T cell factor(s) (ThF). II. Induction of IgG production in B lymphoblastoid cell lines and identification of T cell-replacing factor- (TRF) like factor(s). , 1982, Journal of immunology.

[30]  T. Hirano,et al.  Immortalization of BGDF (BCGF II)- and BCDF-producing T cells by human T cell leukemia virus (HTLV) and characterization of human BGDF (BCGF II). , 1985, Journal of immunology.

[31]  N. Yoshida,et al.  B cell growth factors and B cell differentiation factor from human T hybridomas. Two distinct kinds of B cell growth factor and their synergism in B cell proliferation , 1983, The Journal of experimental medicine.

[32]  T. Hirano,et al.  Phorbol ester increases the level of interleukin 2 mRNA in mitogen-stimulated human lymphocytes. , 1984, Journal of immunology.