Synthesis and evaluation of monoclonal antibody againstPlasmodium falciparum merozoite surface antigen 2

摘要

Objective:To assess the quality of expressedMSP-2 and also to confirm the immune response against different domains of these proteins.Methods:Mice were immunized with a schizont extract to stimulate the immune system to make antibodies against different antigens of the late stage parasite including production of antibodies against different domains of Plasmodium falciparum(P. falciparum)MSP-2.B lymphocytes of immunized mice were extracted from the spleen and the fusion was performed usingNS-1 myeloma cells and the hybridoma cells were assayed byELISA either with a schizont extract or different domains ofMSP-2 and/or byIFAT with whole schizont preparation.Fusion ofNS-1 and spleen cells was performed.The positive hybrids were cloned andELISA was applied against different dilutions.The positive clones were transferred to a small tissue culture flask and after developing they were assayed against schizont extract and the differentMSP-2 domains.The positive clones were expanded to large (75 cm2) flask and cultured under the same conditions, checking them using bothELISA and IFAT and the positive cells were frozen as soon as possible.Results:A total number of7 fusions including26 plates(2496 wells) were performed, ofwhich1336 hybrids were produced and the overall efficiency(1336/2496í100) was about53%.ELISA was performed to detect the positive hybrids against crude schizont extract by which the highest frequency to crude schizont extract was found for the supernatant of the hybrids produced in fusion number3(66 out of315 hybrids). The supernatant of bothB5 andF1 hybridoma cells were more positive against domain2 of the MSP-2 recombinant protein inWestern blotting test.Western blotting results also showed that different domains of theMSP-2 recombinant protein and also theMSP-2 of theP. falciparum parasite were recognized by some of the positive clones and also immune sera.Conclusions:Bringing together all the results of this study it has been confirmed that some clones have recognized both schizont extract and different domains of theMSP-2 recombinant protein and therefore confirming the quality of theMSP-2 domains.