Abstract
A method which allows an accurate determination of very low titers of antibodies able to neutralize diphtheria toxin has been developed. The assay is based on the incubation of a reference amount of diphtheria toxin with different dilution of the serum and the evaluation of the residual toxicity of diphtheria toxin on Vero cells. The cells are seeded in 96-well plastic plates with the toxin-serum mixtures and the residual toxin activity is measured as the block of cell protein synthesis after incubation with [14C]leucine. Antitoxin titers as low as 0.002 IU can be reproducibly and accurately measured and the method gives a linear response in the range 0.002 to 8.0 IU/ml.
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References
Bigio M, Rossi R, Nucci D, Antoni G, Rappuoli R, Ratti G (1987) Conformational changes in diphtheria toxoids. FEBS Lett 218:271–276
Camargo ME, Silveira L, Furuta JA, Oliveira EPT, Germek OA (1984) Immunoenzymatic assay of anti-diphtheric toxin antibodies in human serum. J Clin Microbiol 20:772–774
Cellesi C, Michelangeli C, Rossolini GM, Giovannoni F, Rossolini A (1989a) Immunity to diphtheria, six to 15 years after a basic three-dose immunization schedule. J Biol Stand 17:29–34
Cellesi C, Zanchi A, Michelangeli C, Giovannoni F, Sansoni A, Rossolini GM (1989b) Immunity to diphtheria in a sample of adult population from central Italy. Vaccine 7:417–420
Christenson B (1986) Is diphtheria coming back? Ann Clin Res 18:69–70
Hendriksen CFM, Gun van der JW, Kreeftenber JG (1989) Combined estimation of tetanus and diphtheria antitoxin in human sera by the in vitro toxin-binding inhibition (ToBI) test. J Biol Stand 17:191–200
Kjeldsen K, Simonsen O, Heron I (1985) Immunity against diphtheria 25–30 years after primary vaccination in childhood. Lancet 1:900–902
Kjeldsen K, Simonsen O, Heron I (1988) Immunity against diphtheria and tetanus in the age group 30–70 years. Scand J Infect Dis 20:177–185
Kwantes W (1984) Diphtheria in Europe. J Hyg Camb 93:433–437
Mark A, Christenson B, Granstrom M, Strandell A, Wickborn B, Bottiger M (1989) Immunity and immunization of children against diphtheria in Sweden. Eur J Clin Microbiol Infect Dis 8:214–219
Middlebrook JL, Dorland RB, Leppla SH (1978) Association of diphtheria toxin with Vero cells: demonstration of a receptor. J Biol Chem 253:7325–7330
Miyamura K, Nishio S, Ito A, Murata R, Kono R (1974) Micro cell culture method for determination of diphtheria toxin and antitoxin titers using Vero cells. J Biol Stand 2:189–201
Mortimer J, Melville-Smith M, Sheffield F (1986) Diptheria vaccine for adults. Lancet II: 1182–1183
Naumann P, Hagedorn HJ, Paatz R (1983) Diphtherie-Immunität und ihre epidemiologische Bedeutung. Dtsch Med Wochenschr 108:1090–1096
Pappenheimer AR Jr (1977) Diphtheria Toxin. Annu Rev Biochem 46:69–94
Pappenheimer AR (1981) Diphtheria: studies on the biology of an infectious disease. Harvey Lect 76:45–73
Pappenheimer AR, Uchida T, Harper AA (1972) An immunological study of the diphtheria toxin molecule. Immunochemistry 9:891–906
Pharmacopeia of the USA (1965) Diagnostic diphtheria toxin U.S.P., New York, pp 209
Rappuoli R, Perugini M, Marsili I, Fabbiani S (1983) Rapid purification of diphtheria toxin by phenyl sepharose and DEAE-cellulose chromatography. J Chromatogr 268:548–548
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Padovan, E., Papini, E., Rappuoli, R. et al. Determination of diphtheria toxin neutralizing antibody titers with a cell protein synthesis inhibition assay. Med Microbiol Immunol 180, 29–35 (1991). https://doi.org/10.1007/BF00191698
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DOI: https://doi.org/10.1007/BF00191698