Development of human monoclonal antibodies: A review

This article describes the current status in the development of human monoclonal antibodies. Over the last ten years a lot of information about the human immune system has emerged. Combining these with the many new (bio-)technologies it is plausible that the long awaited breakthrough of this technology is close. This paper focuses on the classical cell-biological methods of achieving stable, antibody-producing human cell lines via cell fusion methods or virus derived transformations of human B-lymphocytes, as well as genetic engineering methods e.g. DNA libraries or phage display technology. The available in vitro immunization methods are critically reviewed and their impact on this topic is discussed. Therapeutic applications for cancer treatment or passive immunization against infectious diseases with antibodies derived by both ways are also reviewed.     

SSGF-I,a potent growth-promoting substance for mammalian cells from swine serum

A small amount of swine serum markedly stimulated cell growth for high productivity subclones derived from a mouse human-human heterohybridoma, N12-16.63, secreting an anti-tetanus toxoid human monoclonal antibody in a polyethylene glycol (PEG)-containing serum-free medium, PEG-86-1. A growth promoting substance, SSGF-I, was isolated from the serum by ammonium sulfate fractionation, Cibacron blue F3A-G affinity chromatography, DEAE-agarose ion exchange chromatography, and gel filtrations on Trisacryl GF 2000 and Sephacryl S-300. SSGF-I was characterized as a low density lipoprotein (LDL) of swine serum by its physico-chemical properties. It promoted cell growth synergistically with PEG and its optimum concentration was 1 to 100g/ml. Human LDL was less active, and human or swine high density lipoprotein (HDL) and very low density lipoprotein (VLDL) were inactive. Based on these results, we propose an improved serum-free medium, PEG-86-3, which contains all the ingredients of PEG-86-1 and 10g/ml SSGF-I. This medium is useful for not only high productivity heterohybridomas but also for a variety of lymphoid cell lines.     

Functional expression of human and mouse low density lipoprotein receptors in hybridomas

Though a mouse.human-human heterohybridoma, N12-16.63, secreting an antitetanus toxoid human monoclonal antibody grew well in a serum-free medium, its high producing subclone N12-69 required SSGF-I, a low density lipoprotein (LDL) from swine serum, or human-LDL (h-LDL) for growth. The growth-promoting action of SSGF-I was caused by its lipid fraction, and SSGF-I could be replaced completely with cholesterol in the presence of bovine serum albumin (BSA). Thus, cell line N12-69 is a cholesterol auxotroph of the heterohybridoma. N12-69 cells express both mouse and human LDL receptors on the cell surface in a ratio of 1:4. SSGF-I bound to both receptors with the same binding affinity, and h-LDL was also take up by the same receptors, though the affinity constant of the receptors for SSGF-I was 1.5 times stronger than that for h-LDL. The growth of N12-69 cells was completely inhibited by the addition of dextran sulfate, which is known to inhibit the binding of LDL to LDL receptors, to an SSGF-I or h-LDL containing medium but was not inhibited at all when dextran sulfate was added to a serum-free medium supplemented with cholesterol and BSA. Furthermore, an anti-human LDL receptor monoclonal antibody partially inhibited the growth of N12-69 cells in an SSGF-I or h-LDL containing medium. These findings suggest that N12-69 cells express both biologically active mouse and human LDL receptors on their cell surfaces and that SSGF-I or h-LDL is taken up by the both receptors to be utilized as a cholesterol source for the growth.