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Low Molecular Weight Factors Displaying Augmenting Activity for Human Antibody Production in vitro
Author(s) -
Haraguchi Soichi,
Hongo Teruaki,
Ishihara Hideki,
Matsuo Tetsumichi,
Yoshida Takato O.
Publication year - 1987
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/j.1348-0421.1987.tb01352.x
Subject(s) - pokeweed mitogen , biology , dialysis tubing , phagocyte , in vitro , sephadex , antibody , microbiology and biotechnology , antigen , cell culture , phagocytosis , immunology , biochemistry , membrane , concanavalin a , enzyme , genetics
Dialyzable low molecular weight antibody‐augmenting factors (LMAAF) were found in the culture supernatant of human tonsillar lymphocytes which were not stimulated by antigen and/or mitogen in vitro. Phagocyte‐depleted nylon wool‐adherent lymphocytes (M − Ny + cells) were responsible for the release of the LMAAF. Marbrook's culture system was adopted to assay for the LMAAF. The M − Ny + cells, which were cultured without antigen and/or without mitogen in the reservoir of Marbrook's diffusion culture vessel, released the LMAAF, which diffused across a dialysis membrane and significantly augmented the pokeweed mitogen (PWM)‐induced plaque‐forming cell (PFC) response of phagocyte‐depleted lymphocytes (M − cells) cultured in the inner vessel. Phagocyte‐depleted nylon wool‐passed lymphocytes (M − Ny − cells) cultured in the reservoir could not augment the PWM‐induced PFC response of the M − cells cultured in the inner vessel. The exuded fluid, which was the dialysate of the culture supernatant of the M − Ny + cells ultrafiltrated with dialysis tubing, also enhanced the PFC response of M − cells cultured in 24‐well multi plates. The exuded fluid also augmented the total IgM and IgG production of human tonsillar and peripheral blood lymphocytes measured by enzyme‐linked immunosorbent assay (ELISA) systems. Gel filtration chromatography on Sephadex G‐25 Superfine column showed that the LMAAF activity was demonstrated in the fractions corresponding to a molecular weight (m.w.) of 362 to 1,355 and a m.w. of 3,560 to 5,700, with a peak activity at about 4,500 dalton. The LMAAF were inactivated by treatment with proteinase K, but not by trypsin, α ‐chymotrypsin, RNase, and DNase, and were stable when treated at 56 C for 60 min. The dialysates of culture supernatants from two out of seven Epstein‐Barr virus (EBV)‐transformed M − Ny + cell lines showed LMAAF‐like activity. These results indicate that phagocyte‐depleted nylon wool‐adherent lymphocytes, possibly B cells, release low molecular weight factors displaying augmenting activity for human antibody production in vitro.