CELLULAR
IMMUNOLOGY
34, 340-353
(1977)
of Murine B Cells by Low Molecular Spec:ific Activation Polyamino-Polycarboxylic Acids (Ampholytes)’ RICHARD C. KNUDSEN,~ AFTAB AHMED,~ ROBERT W. MARTIN VAN CLEAVE, KENNETH W. SELL, AND STEPHEN Clinical
Weight
LONGTON,~
C. GREBE
5
and Experinaental Iwamzmology Department and the Dental Sciences Department, Naval Medical Research Institute, Bethesda, Maryland 20014 Received
July 20,1977
Commercial preparations of ampholytes (AM), which consist of mixtures of large numbers of polyamino-polycarboxylic acids of molecular weight less than 1000 and whose chemical composition is otherwise not specified by the manufacturer, were found to induce blastogenesis in spleen cell cultures from various strains of mice. The blastogenic response of the spleen cells to the ampholytes was 2 to 12 times greater than that of the unstimulated cultures, and peak stimulatory activity occurred 2-4 days after stimulation. Preparations consisting of either acidic, neutral, or alkaline ampholytes were all found to be mitogenic, although the alkaline ampholytes generally induced the highest stimulation and were active over the widest concentration range (0.0880 pg per culture). Studies using spleen cells from nu/nu mice, CBA/N mice, organ distribution studies, and the use of cytotoxic antisera and complement for the depletion of thymus-derived (T) cells or bone marrow-derived (B) cells suggested that ampholytes are mitogenic for murine B cells. These cells arise early in ontogeny, because the ampholytes were found to be as mitogenic for spleen cells from newborn as for adult mice. Further, in concordance with the characteristics of other B-cell mitogens, injection of mice with ampholytes induced polyclonal antibody synthesis. The possibility that blastogenic stimulation was due to a contaminant was ruled out by demonstrating that anti-lipid A-treated, ultrafiltered, and isoelectric-focused ampholytes retained stimulatory activity. The results of these investigations suggest that commercial ampholyte preparations contain an undetermined number of low molecular weight (
IMMUNOLOGY
34, 340-353
(1977)
of Murine B Cells by Low Molecular Spec:ific Activation Polyamino-Polycarboxylic Acids (Ampholytes)’ RICHARD C. KNUDSEN,~ AFTAB AHMED,~ ROBERT W. MARTIN VAN CLEAVE, KENNETH W. SELL, AND STEPHEN Clinical
Weight
LONGTON,~
C. GREBE
5
and Experinaental Iwamzmology Department and the Dental Sciences Department, Naval Medical Research Institute, Bethesda, Maryland 20014 Received
July 20,1977
Commercial preparations of ampholytes (AM), which consist of mixtures of large numbers of polyamino-polycarboxylic acids of molecular weight less than 1000 and whose chemical composition is otherwise not specified by the manufacturer, were found to induce blastogenesis in spleen cell cultures from various strains of mice. The blastogenic response of the spleen cells to the ampholytes was 2 to 12 times greater than that of the unstimulated cultures, and peak stimulatory activity occurred 2-4 days after stimulation. Preparations consisting of either acidic, neutral, or alkaline ampholytes were all found to be mitogenic, although the alkaline ampholytes generally induced the highest stimulation and were active over the widest concentration range (0.0880 pg per culture). Studies using spleen cells from nu/nu mice, CBA/N mice, organ distribution studies, and the use of cytotoxic antisera and complement for the depletion of thymus-derived (T) cells or bone marrow-derived (B) cells suggested that ampholytes are mitogenic for murine B cells. These cells arise early in ontogeny, because the ampholytes were found to be as mitogenic for spleen cells from newborn as for adult mice. Further, in concordance with the characteristics of other B-cell mitogens, injection of mice with ampholytes induced polyclonal antibody synthesis. The possibility that blastogenic stimulation was due to a contaminant was ruled out by demonstrating that anti-lipid A-treated, ultrafiltered, and isoelectric-focused ampholytes retained stimulatory activity. The results of these investigations suggest that commercial ampholyte preparations contain an undetermined number of low molecular weight (
