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:: دوره 3، شماره 7 - ( 11-1394 ) ::
جلد 3 شماره 7 صفحات 18-21 برگشت به فهرست نسخه ها
Purification of human anti-erythropoietin polyclonal antibodies by precipitation and chromatography as an optimized method with potential application in vaccine studies
چکیده:   (712 مشاهده)

Introduction: Polyclonal antibodies are required to be affinity purified. Improved purification methods of polyclonal antibody provide an opportunity to pick the most purified immunoglobulins as a primary or secondary antibody in immunoassays that are included in many vaccine studies. Two common techniques for purifying proteins is salt precipitation and chromatography purification. Our work focuses on purification of polyclonal antibodies against recombinant human erythropoietin (EPO) antigen using these techniques. Methods: A polyclonal antibody was produced by antigen injection with Freund's adjuvant into female albino rabbits. After separation of immunoglobulins using caprylic acid and ammonium sulfate precipitation, selected samples were analyzed by ion exchange chromatography for separation of polyclonal antibody from albumin. The purified proteins were analyzed by SDS-PAGE and antibody was detected by Western Blot analysis and ELISA. Results of immunodiffusion test detected polyclonal antibody production in rabbits. Results: Caprylic acid precipitation was shown to be a more effective purification method than ammonium sulfate. Analysis of protein by spectrophotometer showed 97.6% purity by caprylic acid and 77% purity by ammonium sulfate method. Western Blot and ELISA tests confirmed the presence of antibody against EPO. Conclusion: These findings suggest that caprylic acid can be used as a quality control method in a production facility with minimal cost. On the other hand, ion exchange chromatography is the most common purification method for proteins. Therefore, combination of these techniques may effectively reduce contaminations in antibody purification procedures which may positively affect the interpretations of vaccine efficacies.

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نوع مطالعه: Original article |
دریافت: ۱۳۹۵/۱۲/۲۳ | پذیرش: ۱۳۹۵/۱۲/۲۳ | انتشار: ۱۳۹۵/۱۲/۲۳
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DOI: 10.18869/acadpub.vacres.3.7.18


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Maboudi K, Yaghoubi H, Hadadian S. Purification of human anti-erythropoietin polyclonal antibodies by precipitation and chromatography as an optimized method with potential application in vaccine studies. vacres. 2016; 3 (7) :18-21
URL: http://vacres.pasteur.ac.ir/article-1-92-fa.html
Purification of human anti-erythropoietin polyclonal antibodies by precipitation and chromatography as an optimized method with potential application in vaccine studies. تحقیقات واکسن. 1394; 3 (7) :18-21

URL: http://vacres.pasteur.ac.ir/article-1-92-fa.html

دوره 3، شماره 7 - ( 11-1394 ) برگشت به فهرست نسخه ها
تحقیقات واکسن Vaccine Research
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