Production, purification and HRP conjugation of polyclonal IgG for ELISA test against rabbit Immunoglobulins in Wistar rats

MAIN

©1996-2018. All Rights Reserved. Online Journal of Veterinary Research . You may not store these pages in any form except for your own personal use. All other usage or distribution is illegal under international copyright treaties. Permission to use any of these pages in any other way besides the before mentioned must be gained in writing from the publisher. This article is exclusively copyrighted in its entirety to OJVR. This article may be copied once but may not be, reproduced or re-transmitted without the express permission of the editors. This journal satisfies the refereeing requirements (DEST) for the Higher Education Research Data Collection (Australia). Linking:To link to this page or any pages linking to this page you must link directly to this page only here rather than put up your own page.

OJVRTM

Online Journal of Veterinary Research©

Volume 18(1):12-19, 2014. Redacted 2017.

Production, purification and HRP conjugation of polyclonal IgG for ELISA test against rabbit Immunoglobulins in Wistar rats

Moghadaszadeh M^a*(DVM), Majidi J^b(Ph.D), Abdolalizadeh J^b ( Ph.D), Moghadaszadeh M^c (MD), Askari N^a ( DVM) , Ghaffarilaleh V^d ( Ph.D)

^aShahid Bahonar University of Kerman, Kerman, ^bDrug Applied Research Center, ^cInternal Medicine Department Tabriz, University of Medical Sciences, Tabriz,^dDepartment of Animal Production, Veterinaria Facultad Autonoma, University de Barcelona, Spain.

ABSTRACT

Moghadaszadeh M, Majidi J, Abdolalizadeh J, Moghadaszadeh M, Askari N, Ghaffarilaleh V., Production, purification and HRP conjugation of polyclonal IgG for ELISA test against rabbit Immunoglobulins in Wistar rats, Onl J Vet Res., 18(1):12-19, 2014. Antibodies are used in many diagnostic and experimental procedures. The primary goal in experimental polyclonal antibody production is to obtain sufficient volumes of high titer and high-affinity antibodies economically. Anti-rabbit Immunoglobulins and its conjugate with Horse Radish Peroxides (HRP) is used to diagnose rabbit diseases by ELISA or western blotting tests. In this study, production, purification and HRP conjugation of polyclonal IgG against rabbit Immunoglobulins in rats was performed. Five 6-month-old Male Wister rats were immunized by rabbit Immunoglobulins in combination with Freund’s adjuvant. Purified antibody (using ion-exchange chromatography) was labeled to HRP. Direct enzyme linked Immune-sorbent assay (ELISA) was used to determine the optimum titer and cross reactivity of HRP conjugated IgG. The purity of various IgG preparations was about 98%. The optimum dilution of prepared HRP conjugated IgG was 1:25600. This conjugated IgG has no cross reactivity with guinea pig and hamster Immunoglobulins at optimized dilution. The result of the present study showed that ion-exchange chromatography could be considered as an appropriate method for purification of IgG antibodies.

Key words: Purification, Polyclonal antibody, Horse Radish Peroxides conjugation, Ion exchange chromatography.

MAIN

FULL-TEXT (SUBSCRIPTION OR PURCHASE TITLE)