Abstract
A neutral red assay involving Vero cells was used to quantitate the cytotoxic activity of verotoxins (VT) produced by Escherichia coli and to investigate changes in titer caused by altering the composition of the cell culture medium. Three variations on medium 199 were investigated: one involved supplementing the medium with 5% fetal bovine serum (FBS), a second was the use of serum-free (SF) medium that contained 5% bovine serum albumin and 5 μg of fibronectin per ml, and the third involved the use of 4% Ultroser G, a commercial serum replacement. The level of cytotoxicity varied markedly with the type of VT and with the medium that was used. For VT1, there was no difference in cytotoxicity between medium with 5% FBS and SF medium, but cytotoxicity was reduced more than 100-fold in medium containing Ultroser G compared with cytotoxicity in the other media. For VT2, VT2v, and VTe, there was a slight reduction in cytotoxicity in medium containing 4% Ultroser G and a more marked reduction in SF medium compared with cytotoxicity in medium containing 5% FBS.
Full text
PDF
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Babich H., Borenfreund E. Cytotoxic effects of food additives and pharmaceuticals on cells in culture as determined with the neutral red assay. J Pharm Sci. 1990 Jul;79(7):592–594. doi: 10.1002/jps.2600790709. [DOI] [PubMed] [Google Scholar]
- Gannon V. P., Gyles C. L., Friendship R. W. Characteristics of verotoxigenic Escherichia coli from pigs. Can J Vet Res. 1988 Jul;52(3):331–337. [PMC free article] [PubMed] [Google Scholar]
- Greer C. N., Shewen P. E. Automated colorimetric assay for the detection of Pasteurella haemolytica leucotoxin. Vet Microbiol. 1986 Jun;12(1):33–42. doi: 10.1016/0378-1135(86)90039-8. [DOI] [PubMed] [Google Scholar]
- Head S. C., Karmali M. A., Lingwood C. A. Preparation of VT1 and VT2 hybrid toxins from their purified dissociated subunits. Evidence for B subunit modulation of a subunit function. J Biol Chem. 1991 Feb 25;266(6):3617–3621. [PubMed] [Google Scholar]
- Konowalchuk J., Speirs J. I., Stavric S. Vero response to a cytotoxin of Escherichia coli. Infect Immun. 1977 Dec;18(3):775–779. doi: 10.1128/iai.18.3.775-779.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lingwood C. A., Law H., Richardson S., Petric M., Brunton J. L., De Grandis S., Karmali M. Glycolipid binding of purified and recombinant Escherichia coli produced verotoxin in vitro. J Biol Chem. 1987 Jun 25;262(18):8834–8839. [PubMed] [Google Scholar]
- MacLeod D. L., Gyles C. L. Purification and characterization of an Escherichia coli Shiga-like toxin II variant. Infect Immun. 1990 May;58(5):1232–1239. doi: 10.1128/iai.58.5.1232-1239.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ruoslahti E. Fibronectin and its receptors. Annu Rev Biochem. 1988;57:375–413. doi: 10.1146/annurev.bi.57.070188.002111. [DOI] [PubMed] [Google Scholar]
- Tyrrell G. J., Ramotar K., Toye B., Boyd B., Lingwood C. A., Brunton J. L. Alteration of the carbohydrate binding specificity of verotoxins from Gal alpha 1-4Gal to GalNAc beta 1-3Gal alpha 1-4Gal and vice versa by site-directed mutagenesis of the binding subunit. Proc Natl Acad Sci U S A. 1992 Jan 15;89(2):524–528. doi: 10.1073/pnas.89.2.524. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Valdivieso-Garcia A., Rosendal S., Allen O. B., Thompson C. M., Watson S. Cytotoxicity of Mycoplasma mycoides subspecies mycoides for cultured endothelial cells. Zentralbl Bakteriol. 1989 Dec;272(2):202–209. doi: 10.1016/s0934-8840(89)80008-8. [DOI] [PubMed] [Google Scholar]
- Waddell T., Head S., Petric M., Cohen A., Lingwood C. Globotriosyl ceramide is specifically recognized by the Escherichia coli verocytotoxin 2. Biochem Biophys Res Commun. 1988 Apr 29;152(2):674–679. doi: 10.1016/s0006-291x(88)80091-3. [DOI] [PubMed] [Google Scholar]