Stability Determination of Recombinant Echinococcus Granulosus Antigen B Kit by Physical and Bacteriostatical Methods

Document Type : Original Article (s)

Authors

1 MSc Student, Department of Parasitology and Mycology, School of Medicine AND Student Research Committee, Isfahan University of Medical Sciences, Isfahan, Iran

2 Professor, Cellular and Molecular Biology Research Center AND Department of Biotechnology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran

3 Assistant Professor, Department of Parasitology and Mycology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran

4 Assistant Professor, Cellular and Molecular Biology Research Center AND Department of Biotechnology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran

Abstract

Background: Cystic hydatidosis disease (CHD) is a common disease of human and animal caused by contamination with the metacestode stage of echinococcus granulosus. The disease causes cyst in tissue body of patients. Antigen B is one of the important antigens in liquid cyst.Methods: Expressed and purified recombinant antigen of echinococcus granulosus was used as antigen in ELISA (Enzyme-linked immunosorbent assay) method. The handmade kit was studied with physical and bacteriostatical methods. Physical stability was assessed with accelerated stability test (Arrhenius equation). First, numeral negative and positive samples were evaluated; then, the kit was exposed in special temperatures and times; afterwards, the samples were evaluated again and compared with the first evaluation. Proclin300, Soduim azide and Bacillus subtilis were chosen in bacteriostatic method. Different concentration of bacillus and different percents of preservatives were added to solutions of the kit. Growth of bacteria was evaluated and the least percent of preservatives that had the most effect of bacteriostatic was determinated.Findings: According to stability test, the handmade kit had one-year stability and do not have stability for two years in the 2-8ºC tempreture. Sodium azide had destructive effect on solutions of the kit; but, proclin300 with the concentration of 0.05% was appropriate for solutions.Conclusion: Stability and bacteriostatic methods showed that the handmade kit can be maintained in 2-8ºC with significant activity.

Keywords

References

  1. Athari A, Ansari N, Ourmazdi H, Bijan H, Janbakhsh B, Haghighi A. Essential of medical helminthology-manifestation and treatment of the diseases. 1st ed. Tehran, Iran: Noor Danesh Publication; 2003. [In Persian].
  2. Arfaa F. Medical helmintology. 5th ed. Tehran, Iran: Keshavarz Publication; 2002. [In Persian].
  3. Ghafarifar F, Dalimi-Asl A, Jalosian F. Evaluation of DIG-ELISA for diagnosis of human hydatidosis. Journal of Medical Science: Pathobioligy 2001; 4(3-4): 145-56. [In Persian].
  4. Salami SH, Dalimi A, Madani R. Invention and evaluation of dot ELISA for human hydatidosis diagnosis. Daneshvar Med 1999; 6(24): 37-40. [In Persian].
  5. Thompson RC, Lymbery AJ. Echinococcus and hydatid disease. New York, NY: CABI; 1995. p. 411-63.
  6. Ortona E, Rigano R, Margutti P, Notargiacomo S, Ioppolo S, Vaccari S, et al. Native and recombinant antigens in the immunodiagnosis of human cystic echinococcosis. Parasite Immunol 2000; 22(11): 553-9.
  7. Rigano R, Profumo E, Bruschi F, Carulli G, Azzara A, Ioppolo S, et al. Modulation of human immune response by Echinococcus granulosus antigen B and its possible role in evading host defenses. Infect Immun 2001; 69(1): 288-96.
  8. Abdi J, Kazemi B, Mohebali M, Bandehpour M, Rahimi MT, Rokni MB. Gene cloning, expression and serological evaluation of the 12-kDa antigen-B subunit from Echinococcus granulosus. Ann Trop Med Parasitol 2010; 104(5): 399-407.
  9. Sambrook JJ, Russell DW. Molecular cloning: a laboratory manual. 3rd ed. Long Island, NY: CSHL Press; 2001. p. 1-3.
  10. Promega. Instructions for use of product p2211 and 2551. Technical Bulleint No.253 [Online] 2006. Available from: URL: http://www.yrgene.com/sites/default/files/documents/vector/tb253.pdf
  11. Bandehpour M, Seyed N, Shadnoush M, Pakzad P, Kazemi B. Using recombinant Chlamydia major outer membrane protein (MOMP) in ELISA diagnostic kit. Iran J Biotech 2006; 4(4): 239-44.
  12. Wang Y, Schwarz S, Shen Z, Zhang W, Qi J, Liu Y, et al. Co-location of the multiresistance gene cfr and the novel streptomycin resistance gene aadY on a small plasmid in a porcine Bacillus strain. J Antimicrob Chemother 2012; 67(6): 1547-9.
  13. Vasanthakumari R. Textbook of microbiology. In: Baron EJ, Peterson LR, Finegold SM, Editors. Bailey and Scott's diagnostic microbiology. New Delhi, DL: BI Publications Pvt Ltd; 2007.
  14. Müller R. Worms and human disease. Uganda, Africa: CABI; 2002. p. 85-93.
  15. Mamuti W, Yamasaki H, Sako Y, Nakao M, Xiao N, Nakaya K, et al. Molecular cloning, expression, and serological evaluation of an 8-kilodalton subunit of antigen B from Echinococcus multilocularis. J Clin Microbiol 2004; 42(3): 1082-8.
  16. Sarkari B, Sadjjadi SM, Abidi H, Izadpanah A, Kazemian S, Rafati A. Application of Western Blotting Using Native Antigen B for Serodiagnosis of Human Cystic Echinococcosis. Iranian Journal of Parasitology 2007; 2(3): 7-12.
  17. Kalantari E, Bandehpour M, Pazoki R, Taghipoor-Lailabadi N, Khazan H, Mosaffa N, et al. Application of recombinant Echinococcus granulosus antigen B to ELISA kits for diagnosing hydatidosis. Parasitol Res 2010; 106(4): 847-51.
  18. Magari RT, Murphy KP, Fernandez T. Accelerated stability model for predicting shelf-life. J Clin Lab Anal 2002; 16(5): 221-6.
  19. De Vore K. Have more confidence in your stability data: two points to consider. J Pharm Biomed Anal 2006; 41(1): 293-8.
  20. Hornback LA. Stability testing for IVDs [Online] 2004; Available from: URL: http://www.ivdtechnology.com/article/stability-testing-ivds/
  21. Morris JW. A comparison of linear and exponential models for drug expiry estimation. J Biopharm Stat 1992; 2(1): 83-90.
Journal of Isfahan Medical School
Volume 31, Issue 237 - Serial Number 237
March and April 2013
Pages 701-711

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History

  • Receive Date: 16 August 2012
  • Revise Date: 01 November 2024
  • Accept Date: 06 April 2022

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