Natural acquired inhibitory antibodies to Plasmodium vivax Duffy binding protein (PvDBP-II) equally block erythrocyte binding of homologous and heterologous expressed PvDBP-II on the surface of COS-7 cells

Abstract

The binding domain of Plasmodium vivax Duffy binding protein (PvDBP-II) is a promising blood-stage vaccine candidate for vivax malaria. For the development of a successful vivax malaria vaccine based on DBP-II, the antigenic diversity and also naturally occurring functional antibodies to different PvDBP-II variant types in the various populations must be determined. However, similar to other blood-stage antigens, allelic variation within the PvDBP-II is a fundamental challenge for the development of a broadly efficient vaccine. The present study was performed to define whether the polymorphisms in PvDBP-II influence the nature of functional inhibitory activity of naturally acquired or induced anti-DBP-II antibodies in mice. In this investigation, five genetically distinct variants of PvDBP-II were transiently expressed on the COS-7 cell surface. Erythrocyte-binding inhibition assay (EBIA) was performed using human sera infected with corresponding and non-corresponding P. vivax variants as well as by the use of mice sera immunized with different expressed recombinant PvDBP-IIs. EBIA results showed that the inhibitory percentage varied between 50 and 63 % by using sera from infected individuals, and in case of mouse antisera, inhibition was in the range of 76-86 %. Interestingly, no significant difference was detected in red blood cell binding inhibition when different PvDBP-II variants on the COS-7 cell surfaces were incubated with heterologous and homologous sera infected with PvDBP-II variants. This suggests that the detected polymorphisms in all five forms of PvDBP-II may not affect functional activity of anti-DBP-II antibodies. In conclusion, our results revealed that there are functional cross-reactive antibody responses to heterologous PvDBP-II variants that might provide a broader inhibitory response against all, or at least the majority of strains compared to single allele of this protein that should be considered in development of PvDBP-II-based vaccine.

Th1 immune response to Plasmodium falciparum recombinant thrombospondin‐related adhesive protein (TRAP) antigen is enhanced by TLR3‐specific adjuvant, poly(I:C) in BALB/c mice

“…No binding of rPvDBP (unrelated recombinant protein) to HepG2 liver cells was observed. rPvDBP was expressed in E. coli and has been used for mouse immunization with CFA / IFA as adjuvant . rPvDBP (as unrelated antigen) and mouse anti‐ rPvDBP antibody were used to test the binding of rPvDBP to HepG2 liver cells (negative control).…”

“…E. coli expressed rPvDBP was used in similar concentrations with rPfTRAP. After 3 times washing with TBS, the wells were incubated with polyclonal antibodies (1:200 dilution) raised in mice against purified rPfTRAP or rPvDBP as appropriate at RT for 90 min, followed by incubation with goat anti‐mouse IgG‐horseradish peroxidase (HRP) conjugate (1:25 000; Sigma‐Aldrich) at RT for 60 min. The reaction was developed using 3,3′,5,5′‐tetramethylbenzidine (TMB; Sigma, USA) as substrate and binding of rPfTRAP and rPvDBP (unrelated recombinant protein as control) to HepG2 cells was measured at 450 nm using a microplate reader (Biotek, Winooski, VT, USA).…”

“…The reaction was developed by adding the TMB (Sigma‐Aldrich) as substrate and then was stopped by 2 N H 2 SO 4 , and the absorbance was measured at 450 nm. To evaluate the specificity of IgG antibody responses to rPfTRAP, recombinant proteins (rPvDBP and rPfTRAP) were used in ELISA with unrelated antibodies (anti‐rPfTRAP and anti‐rPvDBP, respectively). Anti‐rPfTRAP IgG subclasses were detected as described above in exception of incubation of 1:1000 dilution of goat anti‐mouse IgG1, IgG2a, IgG2b and IgG3 antibodies (Sigma‐Aldrich) at RT for 1 h as secondary antibodies.…”

Genetic polymorphism of Plasmodium vivax Duffy Binding Protein in malarious areas in southeastern of Iran

“…The relevant gene contains some diversities depending on different geographical regions (Babaeekho et al 2009), nevertheless the gene does not bear more polymorphism in comparison with merozoite surface protein (MSP) and circumsprozoite protein (CSP) genes. Therefore PvDBP is supposed to be a trustable candidate for P. vivax vaccine (Premaratne et al 2011;Valizadeh et al 2016). Learning about polymorphism of PvDBP gene guides us to understand which allele is more important as a specific target to produce a vaccine.…”

Unusual mtDNA Control Region Length Heteroplasmy in the COS-7 Cell Line

“…The resulting cell line retains complete permissiveness for the lytic growth of SV40, supports the replication of tsA209 virus at 40 degrees C and supports the replication of pure populations of SV40 mutants with deletions in the early region, which is consistent with the integration of the early region of SV40, including wild-type t-antigens. It was originally developed for the propagation of pure populations of recombinant SV40 viruses; however, its uses also includes the propagation of polyomavirus and rotavirus [3,4], as well as research on ion channels [5], apoptosis and autophagy [6], immunology [7], cell biology [8], vitamin [9], hormone [10] and cell signaling [11,12].…”