medigraphic.com
ISSN 1025-0298 (Electronic)
ISSN 1025-028X (Print)

2014, Number 2

<< Back Next >>

VacciMonitor 2014; 23 (2)

Using the caprylic acid in obtaining the horse immunoglobulin anti tetanus toxin

Miranda-Cruz AR, Sánchez-Artigas R, Otero-Alfaro O, Góngora-Amores W, Cobos-Valdes D, Goya-Batista Y, Balboa-González J, Pérez-Martín O

Language: Spanish
References: 20
Page: 63-72
PDF size: 202.64 Kb.


ABSTRACT

Purified horse anti tetanus toxin immunoglobulins with high antibody concentration and biological activity, are used as heterologous sera and standard serum for corresponding vaccine. However, the selection of precipitating agents for the purification of them is a problem in terms of purity, yield and economy, which still remains to be solved. In this study, a procedure to fractionation horse plasma anti tetanus toxin by Caprylic acid precipitation is described. A solution of acetic acid/sodium acetate was added to the samples to achieve different ionic strengths, 0.05-0.4 moles/L, and caprylic acid was added to each volume to reach final concentrations from 1 to 7% (v / v), with vigorous stirring for 60 min. Non-immunoglobulin proteins precipitated under these conditions, while immunoglobulins remained in the supernatant, which was then diafiltered to remove caprylic acid and concentrate inmunoglobulins. This methodology was compared to that based on ammonium sulfate fractionation. The best results were given when caprylic acid was added to plasma, until a final caprylic acid concentration of 3% was reached, at ionic strength 0.2 moles/L and pH adjusted to 4.5. The IgG recovery was 91-95% by using the caprylic acid method. It was achieved a protein concentration of 27.1-29.3 mg/mL and an albumin/immunoglobulins ratio of 0.019-0.021. The purity was 91- 95% and the anti-tetanus toxin activity recovery of 93-96%.


REFERENCES

  1. Sjostrom L, Al-Abdulla I, Rawat S, Smith D, Landon J. A comparison of ovine and equine antivenoms. Toxicon1994;32:427-33.

  2. Morais VM, Massaldi H. Snake antivenoms: adverse reactions and production technology. J Venom Anim Toxins Trop Dis 2009;15(1):2-18.

  3. Wagner KS, Stickings P, White JM, Neal S, Crowcroft NS, Sesardic D, et al. A review of the international issues surrounding the availability and demand for diphtheria antitoxin for therapeutic use. Vaccine 2010;28:14-20.

  4. Raweerith R, Ratanabanangkoon K. Fractionation of equine antivenom using caprylic acid precipitation in combination with cationic ion-exchange chromatography. J Immunol Meth 2003;282:63-72.

  5. Steimbuch M, Audran R. The isolation of IgG from mammalian sera with the aid of caprylic acid. Arch Biochem Biophys 1969;134:279-84.

  6. Chen S, Lau H, Brodsky Y, Kleemann GR, Latypov RF. The use of native cation-exchange chromatography to study aggregation and phase separation of monoclonal antibodies. Protein Sci2010;19(6):1191-1204.

  7. Guang-Ping R, Li M, Xiao-Wei H, Min-Jie M, Yong Z, Ming-Qian Z, et al. Efficient production, purification, and application of egg yolk antibodies against human HLA-A*0201 heavy chain and light chain (β2m). Protein Expr Purif 2005;44:45-51.

  8. Theakston RDG, Warrell DA, Griffiths E. Report of a WHO workshop on the standardization and control of antivenoms. Toxicon 2003;41:541-57.

  9. León G, Lomonte B, Gutiérrez JM. Anticomplementary activity of equine whole IgG antivenoms: comparison of three fractionation protocols. Toxicon 2005;45:123-8.

  10. Morais V, Massaldi H. Effect of pepsin digestion on the antivenom activity of equine immunoglobulins. Toxicon 2005;46:876-82.

  11. Rojas G, Jiménez JM, Gutiérrez JM. Caprylic acid fractionation of hyperimmune horse plasma: description of a simple procedure for antivenom production. Toxicon 1994;32:351-63.

  12. Nudel BC, Perdoménico C, Iácono R, Cascone O. Optimization by factorial analysis of caprylic acid precipitation of nonimmunoglobulins from hyperimmune equine plasma for antivenom preparation. Toxicon 2012;59:68-73.

  13. Morais V, Massald H. A model mechanism for protein precipitation by caprylic acid: Application to plasma purification. Biotechnol Appl Biochem 2012;59(1):50-4.

  14. Miranda AR, Sánchez R, Góngora W, Mulet O. Obtención de sueros hiperinmunes de origen equino antitoxina tetánica, utilizando adyuvante oleoso. COCMED 2006;10(3). Disponible en: http://www.cocmed.sld.cu/no103/ n103ori8.htm

  15. Lowry DH, Rosembroug NJ, Farr AL, Randal R. Protein measurement with foling phenol reagent. J Biol Chemical 1951;193:256-69.

  16. Doumas BT, Watson WA, Biggs HG. Albumin Standards and the Measurement of Serum Albumin with Bromocresol Green. Clin Chim Acta 1971;31:87-96.

  17. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970;227(5259):680-5.

  18. Vojdani F. Solubility. In: Methods of Testing Protein Functionality. GM Hall, ed. New York: Blackie Academic & Professional; 1996.p.11-60.

  19. Gutiérrez JM, Rojas E, Quesada L, León G, Núñez J, Laing GD, et al. Pan-African polyspecific antivenom produced by caprylic acid purification of horse IgG: an alternative to the antivenom crisis in Africa. Trans R Soc Trop Med Hyg 2005;99:468-75.

  20. Wang J, Diehl T, Aguilar D, Dai XP, Arunakumari A. Precipitation of process-derived impurities in non-protein a purification schemes for antibodies. BioPharm International 2009;10(Suppl 3):4-10.




2020     |     www.medigraphic.com