Preparation And Identification Of Monoclonal Antibody Against GP2 Of Lymphocytic Choriomeningitis Virus

Backround:Lymphocytic choriomeningitis virus(LCMV)belongs to arenavirus family,and rodents is its host in nature,such as house mouse,pet mouse and hamster.It can transmit through saliva,nasal excretion,milky juice,semen,urine and feces,it,as well,can transmit through other approaches,such as the wound of skin or mocusa,placenta and aerosol in air.Most of the host appear no symptom when they are infected,which provides a premise for it to transmit wildly and unperceivably.So we usually underestimate the universality of LCMV infection worldwide.We have obtained milestone-like discoveries in restriction of MHC molecule on T cell,T cell exhaustion and immunotolerance by means of LCMV research.Among LCMV,the subtype virus Armstrong and Cl-13 have tiny variation in genome,but they can lead to significant difference of acute or chronic infection in host,so they are chosen as model for intensive research of pathology in acute or chronic infection.At present,the knowledge of chronic infection from LCMV are extended in many fields such as HIV,tuberculosis,HBV,HCV,autoimmune diseases and tumor.On the other hand,LCMV can infect human through many ways.As human who is infected LCMV seldom appears symptom,or only has a set of mild non-specific symptoms which can self-heal in several days,so we ignore its existence for a long time.But It can cause infection by chance under some conditions such as pregnancy,immunodeficiency and the period of solid organ transplantation,so it could bring great menace to life.GP2,as a transmembrane protein,locating on the surface of LCMV particle,decorate the surface of virus with GP1.Moreover,it afford more important function of triggering membrane fusion between virus and cell by transformation when infection occurs.However,researchers still utilize immunized serum or RT-PCR to detect LCMV all over the world,and even to these days,there is no commodity of LCMV GP2 mAb.So it is a significant implication to develop a kind of mAb against GP2 of LCMV.Objective:The purpose of this research is to prepare mAb against GP2 of LCMV,which could be a foundation for preparing enzyme conjunction or fluorescence conjunction testing antibodies hereafter,and could provide possibility for immunotherapy in the future.It applies,as well,for GP2 purification by affinity chromatography,which provides a nice substantial preparing for studying structure and function of GP2,and for exploitation of vaccine against LCMV in further steps.Method:We utilized the technical of cell hybridization to obtain hybridoma,then collected the hybridoma medium.We identified the subclass of the m Ab and the immune potency via immunofluorescence(IFA),dot-blot and ELISA.Subsequently.The mAb was purified by protein G Sepharose affinity chromatography protocol,and then purity of the mAb was checked by SDS-PAGE.At last,Western-blot was performed to detect the site where the m Ab interacted with the LCMV antigen.result:We obtained a hybridoma strain which could secreted specific anti-LCMV m Ab stably,then a kind of purified m Ab was collected favorably.The mAb was demonstrated that it could recognize the antigen from LCMV by IFA and dot-blot.We identified the subclass of the mAb was IgG2 b,and the immune potency of the medium was 1.35×103.SDS-PAGE demonstrated that the process of purifying m Ab through protein G Sepharose affinity chromatography protocol was successfully,the purity of m Ab was high,Western-blot,as well,implied that the site where the mAb combined specifically was GP2 on surface of the LCMV particle.conclusion:We have prepared a kind of high affinity,specific mAb against GP2 of LCMV,which belongs to IgG2 b subclass.