Volume 8, Issue 4 (10-2020)                   JoMMID 2020, 8(4): 155-160 | Back to browse issues page

XML Print

Download citation:
BibTeX | RIS | EndNote | Medlars | ProCite | Reference Manager | RefWorks
Send citation to:

Panahi Z, Kia V, Moghiman M, Doroud D, Shokri R, paryan M. A Fast and Straightforward Method for the Purification of Anti- Immunoglobulin G (IgG) for Coombs Wright Assay. JoMMID. 2020; 8 (4) :155-160
URL: http://jommid.pasteur.ac.ir/article-1-313-en.html
Department of Research and Development, Production and Research Complex, Pasteur Institute of Iran, Tehran, Iran
Abstract:   (73 Views)
Introduction: Immunoglobulin G is the most abundant immunoglobulin in human blood, comprising approximately 75% of serum antibodies. Human IgG is a glycoprotein and can be an antigen for other mammals. Antibody-sensitized red blood cells (RBCs) and complement components can react with anti-human globulins resulting in their agglutination. This study aimed to prepare an antiserum against human IgG for deploying in Coombs Wright and cross-match for rare blood groups. Methods: After isolation of serum from healthy blood donors, serum proteins were precipitated using ammonium sulfate. Consequently, tangential flow filtration and ion-exchange chromatography were applied to purify IgG. SDS-PAGE and Bradford protein content assay was conducted to evaluate the quality and the concentration of the purified IgG.  Rabbits were weekly injected with different amounts of the protein four times. Then, sera were obtained from the immunized mice, and total IgG was purified.  Finally, the Coombs Wright test was performed on samples from brucellosis patients to validate purified IgG antibody quality. Results: Electrophoresis and Bradford assay results showed that the purified protein had considerable high purity and quantity. Protein bands of reducing and the non-reducing SDS-PAGE showed high purity of the protein along with a protein yield of 2.2 mg/L. Coombs Wright tests using the rabbit anti-human serum had a comparable result with available commercial anti-human immunoglobulin. Conclusion: The results indicated that our method for the purification of IgG was suitable for anti-human globulin preparation. This antibody can also be used in clinical diagnostic tests such as Coombs Wright, cross-match, and blood types evaluation with weak Rh or Du antigens. 
Full-Text [PDF 959 kb]   (19 Downloads)    
Type of Study: Original article | Subject: Diagnostic/screening methods and protocols
Received: 2020/11/1 | Accepted: 2020/10/19 | Published: 2021/02/13

1. Eckman MR. Brucellosis Linked to Mexican Cheese. JAMA. 1975; 232 (6): 636-7. [DOI:10.1001/jama.1975.03250060032018]
2. Young EJ. An Overview of Human Brucellosis. Clin Infect Dis. 1995; 21 (2): 283-9. [DOI:10.1093/clinids/21.2.283]
3. Mantur BG. Childhood Brucellosis--a Microbiological, Epidemiological and Clinical Study. J Trop Pediatr. 2004; 50 (3): 153-7. [DOI:10.1093/tropej/50.3.153]
4. Edward J. Young MJC. Brucellosis: Clinical and Laboratory Aspects. 2020.
5. Mantur BG, Biradar MS, Bidri RC, Mulimani MS, K. V, Kariholu P, et al. Protean clinical manifestations and diagnostic challenges of human brucellosis in adults: 16 years' experience in an endemic area. J Med Microbiol. 2006; 55 (7): 897-903. [DOI:10.1099/jmm.0.46097-0]
6. Tsolia M, Drakonaki S, Messaritaki A, Farmakakis T, Kostaki M, Tsapra H, et al. Clinical Features, Complications and Treatment Outcome of Childhood Brucellosis in Central Greece. J Infect. 2002; 44 (4): 257-62. [DOI:10.1053/jinf.2002.1000]
7. Young EJ. Serologic Diagnosis of Human Brucellosis: Analysis of 214 Cases by Agglutination Tests and Review of the Literature. Rev Infect Dis. 1991; 13 (3): 359-72. [DOI:10.1093/clinids/13.3.359]
8. Almuneef M, Memish ZA. Persistence of Brucella Antibodies after Successful Treatment of Acute Brucellosis in an Area of Endemicity. J Clin Microbiol. 2002; 40 (6): 2313. [DOI:10.1128/JCM.40.6.2313.2002]
9. Gazapo E, Lahoz JG, Subiza JL, Baquero M, Gil J, de la Concha EG. Changes in IgM and IgG Antibody Concentrations in Brucellosis Over Time: Importance for Diagnosis and Follow-Up. J Infect Dis. 1989; 159 (2); 219-25. [DOI:10.1093/infdis/159.2.219]
10. Evans RS, Turner E, Bingham M. Chronic Hemolytic Anemia due to Cold Agglutinins: the Mechanism of Resistance of Red Cells to C′ Hemolysis by Cold Agglutinins. J Clin Invest. 1967; 46 (9): 1461-74. [DOI:10.1172/JCI105638]
11. S. Al Dahouk HTKNHN and DF. Laboratory-based diagnosis of brucellosis--a review of the literature. Part II: serological tests for brucellosis. Clin Lab. 2003; 49 (11-12): 577-89.
12. Sanborn, Warren R, Claus Chr. Heuck, Raja El Aouad WBS. Fluorescence microscopy for disease diagnosis and environmental monitoring [Internet]. World Health Organization Regional Office for the Eastern Mediterranean Cairo; Available from: https://apps.who.int/iris/bitstream/handle/10665/119734/dsa281.pdf?sequence=1&isAllowed=y
13. T. J. Kindt RAG and BAO. Kuby immunology. Sixth edition. 2007.
14. Vahdat K, Jafary S M HSM. Seroepidemiological prevalence of brucellosis in livestock breeders of the central rural area of Bushehr province 2003-4. Iran South Med J. 2006; 9 (1): 51-8.
15. Amirzargar AA, Hassibi M, Maleknejad P, Piri-Dougahe H, Jafari S, Soud Bakhsh AR, et al. Comparison of Diagnostic Methods in Hospitalized Patients With Brucellosis in Iran. Infect Dis Clin Pract. 2009; 17 (4): 239-42. [DOI:10.1097/IPC.0b013e31818718e8]
16. Nielsen K. Diagnosis of brucellosis by serology. Vet Microbiol. 2002; 90 (1-4): 447-59. [DOI:10.1016/S0378-1135(02)00229-8]
17. Chegeni AS, Ezatpour B, Saki M, Mokhayeri H, Adavi S, Nasiri E, et al. Seroepidemiology of human brucellosis in nomads in a rural area of Iran. Asian Pacific J Trop Dis. 2014; 4 (4): 333-6. [DOI:10.1016/S2222-1808(14)60584-3]
18. Gómez MC, Nieto JA, Rosa C, Geijo P, Escribano MA, Muñoz A, et al. Evaluation of Seven Tests for Diagnosis of Human Brucellosis in an Area Where the Disease Is Endemic. Clin Vaccine Immunol. 2008; 15 (6): 1031-3. [DOI:10.1128/CVI.00424-07]
19. Kerr MA, Loomes LM. Properties of Immunoglobulins. In: Immunochemistry Labfax. Elsevier; 1994. [DOI:10.1016/B978-0-12-404940-6.50008-6]
20. J. M. Walker. Protein Protocols Handbook. 2002. [DOI:10.1385/1592591698]

Add your comments about this article : Your username or Email:

Send email to the article author

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.