جستجوی مقالات مرتبط با کلیدواژه « Epitope » در نشریات گروه « پزشکی »

Leishmania is a vector-borne protozoon, which causes visceral, cutaneous and mucocutaneous leishmaniosis in human and animals. Monocyte- derived exosome vaccines can be used as prophylaxis and immunotherapy strategies. The aim of this study was to design a multiple-epitope candidate vaccine using leishmaniolysin (GP63) and rK39 proteins against Leishmania major and L. infantum for monocyte-derived exosome preparation.

This study was carried out in Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran, 2023–2024. Effective immunodominant epitopes were selected from two antigenic proteins of GP63 and rK39 using various immunoinformatics and bioinformatics approaches. Vibrio cholerae β- subunit was used as an adjuvant to stimulate immune responses. Then, appropriate linkers were selected for the fusion of epitopes. The 3D model of candidate vaccine was predicted and validated.

This designed candidate vaccine could effectively be used as a prophylaxis strategy against leishmaniosis.

A candidate vaccine was designed using bioinformatic and immunoinformatic studies with virtual acceptable quality; however, effectiveness of this vaccine should be verified through further in-vitro and in-vivo studies.

Hepatitis C Virus infects more than 170 million people globally despite highly effective direct acting antiviral drugs that greatly improved treatment. The Hepatitis C virus envelope glycoproteins E1 and E2 are the major target to induce immune responses. Since, the different aspects of E1 such as its function and structure are still discussed and require further study, in current study critical regions of E1 were evaluated.

Mutation diversity in these areas was determined using strains that were available in online databanks and authentic software. Furthermore, RT-PCR for E1 was done on HCV-1a positive samples and the sequences were analyzed.  The percentage of substitutions, desired and stable residues for mutation in each position were indicated.

The integrated results exhibited bNAb epitope (residues 313-328) which is the most conserved epitope in E1 glycoprotein sequence among all genotypes of HCV.

These kinds of studies may shed light on identification more binding sites of virus and broadly cross-neutralization of antibodies. Moreover, it may facilitate the modeling of peptides to new antiviral design or boosting the immune response in multi-epitope vaccine studies.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread worldwide as an Omicron variant (B.1.1.529). Compared with the original SARS-CoV-2, this variant has more than 30 mutations on its spike. The Lambda variant (known as SARS-CoV-2 lineage C.37) is another variant of interest. The Lambda spike protein bears seven mutations; G75V, T76I, L452Q, F490S, D614G, T859N, and ∆247-253. The effect of such mutations on immune escape from neutralizing antibodies and infectivity is unknown.

In-silico tools were applied to predict the antigenicity of the spikes of Lambda and Omicron variants and the results were compared to the reference Wuhan spike protein antigenicity. SWISS-MODEL, MolProbity, and QMEAN were used for model quality assessment. DiscoTope2.0, BEpro, and Ellipro were used for the prediction of conformational and linear B cell epitopes.

The evaluation of the obtained modeled proteins showed that the predicted models by Swiss-Model had higher quality for Lambda and Omicron spikes with 0.56% and 1.63% of residues in outlier and 94.39% and 92.51% residues in favored regions, respectively. The results of conformational B cell epitope prediction showed 6 epitopic regions on S1 of Lambda spike and 1 epitopic region on the S2 segment of the protein. For the Omicron variant, 9 epitopic regions existed on S1 and 1 epitopic region (1137-1159) was on S2.

Our results suggested that B cell epitope removal and reducing the antigenicity properties of the epitopic residues involve reducing susceptibility to antibody neutralization of the mutant protein

Antibodies, well-known as immunoglobulins (Igs), are produced by B lymphocytes and specifically defend against pathogens. Igs are glycoproteins and have high diagnostic value in several diseases including infections (1). Igs are composed of light and heavy chains (2, 3). Each chain is comprised of about 110-120 amino acid residues which create immunoglobulin folds named domains (4, 5).  Domains play an important role in Igs biological functions. Every domain is included of two anti-parallel β-sheets. B-sheets play an important role in epitope formation because most of the epitopes are located in B-sheets (6, 7).  Epitope/ antigenic determinant is a part of antigen recognized by antibody (8).  For precise detections of Igs, diagnostic tools such as Igs- epitope specific monoclonal antibodies (MAbs) are needed (11). Therefore B-sheets are valuable tools in detection of proteins epitopes.IgG is the highest Ig in serum and has vital role in defense against infections (12). For optimizing current IgG diagnostic tests, accurate determination of IgG-specific epitopes is extremely important (18). Immunoinformatic is a branch of immunology uses a large amount of computer-generated biological information to solve immunological problems and diagnosis of diseases faster and more accurate (19-21). Immunoinformatic has spread to almost all areas of immunology and has exceedingly facilitated understanding of immune responses and their role in disease and health. Accordingly immunoinformatic has provided novel research opportunities to study the molecular mechanisms of immunological processes and related diseases and thus provide more effective diagnostic and treatment strategies (24-26). As B-sheets play an important role in epitope formation (6, 7) and immunogenic epitopes are very useful tools for production of specific monoclonal antibodies against a molecule (11), we conducted this research for recognition of human IgG B-sheets with immunoinformatic method and study their compatibility with IgG epitopes.

The confirmed amino acid sequence of reference human IgG was determined with PDB data bank in http://www.rcsb.org/pdb/home/home.do web site. The second structure of human IgG was obtained with Phyre2 (Protein Homology/analogy Recognition Engine V 2.0) software available on the http://www.sbg.bio.ic.ac.uk/phyre website. The location of B-sheets and epitopes in IgG was determined by the IEDB (Immune Epitope Database) at www.immuneepitope.org. Website.

Figures 1 and 2 show the second structure of light and heavy chains of the human IgG molecule, respectively, obtained by phyre2 software. The IgG light and heavy chains second structure consists of alpha spirals and B-sheets.
Figures 3 and 4 show the location of B-sheets in light and heavy chains of the human IgG molecule obtained by IEDB software.Most of  the  B-sheets in IgG molecules were located in 150 - 175 amino acid sequences of light chains and in 130-140, 160-170, 190-200, 220-245, 375-410 and 420-425 amino acid sequences of heavy chains respectively as was determined by IEDB software.Three epitopes sited to constant domain of IgG light chain (CL) were predicted. These epitopes were located at 150-175 and 180-205 amino acid sequences of light chain. Also three epitopes situated to second and third constant domains of IgG heavy chain (CH2 and CH3) were predicted. These epitopes were located at 200-270, 290-360 and 380-400 amino acid sequences of heavy chain.Based on the results of present study, 66% of human IgG light chain epitopes and 66% of human IgG heavy chain epitopes are situated at the IgG B-sheets locations. Also in present study, 91% of the amino acids forming B-sheets sited in the heavy chains of IgG molecule were located in the CH2 and CH3 domains and 100% of the human IgG heavy chain epitopes were located in the CH2 and CH3 domains

According to the results of this study, most of human IgG epitopes are located in regions of the molecule where B-sheets are most likely situated, indicating a relatively high compatibility of B-sheets with IgG epitopes.Given that the B-sheets present in the second structure of proteins play an important role in epitope formation (6, 7), the occurrence of most human IgG epitopes (66%) at the location of B-sheets seems evident.Moreover, according to the findings of the present study, a total of 91% of the amino acids forming B-sheets sited in the heavy chains of IgG molecule were located in the CH2 and CH3 domains and 100% of the human IgG heavy chain epitopes were located in the CH2 and CH3 domains. This again indicates the high compatibility of B-sheets with epitopes located on the heavy chain of the human IgG molecule.The findings of our study are confirmed by Hajighasemi et al. study (31). In study of Hajighasemi et al. (31), about 91% of the linear epitopes located on the heavy chains of the IgG molecule, were situated in the CH2 and CH3 domains. Similarly, in our study 100%  of human IgG heavy chain  epitopes located on CH2 and CH3 domains.Also the results of present study are confirmed by another research (36) in which all (100% ) of spatial epitopes on fragment of crystallizable [Fc] component (part of heavy chains)  of the human IgG molecule were located in CH3 and CH2 domains as were  identified by DiscoTope and ElliPro  immunoinformatic softwares. Similarly in the present study, 100%  of IgG heavy chain  epitopes were located in the CH2 and CH3 domains.Moreover the results of another study performed by Rohani et al. (37) support the findings of current study. Rohani et al., reported that most of the areas with highest surface accessibility, hydrophilicity and flexibility, were situated in CH2 and CH3 domains of  human IgG heavy chains. (37). According to our results, most of  B-sheets are located in the CH2 and CH3 domains of  human IgG heavy chains. As B-sheets are regions with high accessibility, hydrophilicity and flexibility (6), presence of most B-sheets in the CH2 and CH3 domains  which are reported to be  highly accessible, hydrophilic and flexible ( 37), is reasonable.Furthermore, the high immunogenicity of B-sheets (39), once again confirms results of the present study. As immunogenicity plays an important role in epitope development, the high adaptation between the location of B-sheets and epitopes ( shown in our study) which both are highly immunogenic, seems logical.Besides since B-sheets play an important role in epitope formation(6, 7),  the high compatibility between location of B-sheets and IgG epitopes ( shown in present study) seems sensible.Overall, presence the most of B-sheets and IgG epitopes in CL, CH2 and CH3 domains of human IgG, shows the high compatibility between location of B-sheets and epitopes in human IgG. These findings are precious tools not only for prediction of IgGimmunogenic epitopes in order to producing specific monoclonal anti IgG antibodies to optimize current human IgG diagnostic kits but also for more exact detection of IgG functions. Moreover our results are helpful in producing of similar proteins for diagnostic and therapeutic aims, structure- function relationships and phylogenic studies.

The COVID-19 pandemic is a red alarm for global health, so researchers around the world are working on it to design an effective vaccine against it. Protein is one of the candidates for vaccine development which plays an important role in virus pathogenesis. Accordingly, this study was done to evaluate the critical characteristic of this protein as a vaccine candidate using in-silico analysis. The sequence of SARS-CoV-2-associated E protein was recruited from NCBI and subjected to the IEDB software to evaluate the most potent epitopes. The capacity of the interactions of HLA-I and HLA-II molecules with selective peptides was studied using IEBD tool kit. The E protein sequence was subjected to B cell and T cell tests to realize the most promising peptides that could act as COVID-19 vaccine. Among the tested peptides for the T cell-test, this study found two interesting epitopes: VSEETGTLI and LTALRLCAY that exhibit high binding affinity as a strong indicator to HLA-I and HLA-II alleles together. The results of the analysis demonstrated that some epitopes in the E protein have a relatively higher immunogenicity score based on interaction with HLA-II, such as SEETGTLIVNSVLLF, TLIVNSVLLFLAFVV, LAFVVFLLVTLAILT, LAILTALRLCAYCCN, and SVLLFLAFVVFLLVT. Furthermore, two sequences (FVSEET and PSFYVYSRVKNLNSSRVP) were reported as the selective linear epitopes for B cell, on the surface of SARS-CoV-2 E protein and being Immunogenic. Since E protein can stimulate favorable immune responses, T and B- cell responses, its evaluation in patients with COVID-19 is of a great importance.

Due to high genetic variation in human leukocyte antigen )HLA( alleles, epitope-based vaccines don’t show equal efficacy in different human populations. therefore, we proposed a multi-epitope vaccine against SARS-CoV-2 for Iranian populations.

For this purpose, the proteins without allergenicity and high antigenicity as well as conservancy level from SARS-CoV-2 were subjected to in silico B- and T-cell epitope mapping. T-cell epitope mapping process was performed based on the most frequent human leukocyte antigen (HLA) alleles in Iran. The B- and T-cell epitopes were screened based on their allergenicity, antigenicity and hemolytic potential and the epitopes with appropriate properties were subjected for final vaccine construct. The screened B- and T-cell epitopes were organized in the final vaccine construct. The secondary and tertiary structures of the proposed vaccine were predicted and its affinity to HLA-I, HLA-II, toll like receptor (TLR-3) and TLR-4 by molecular docking method. Additionally, probable immune responses against the vaccine were predicted trough immune simulation.

The final vaccine construct was includes six, eight and six linear B-cell epitopes, HLA-I restricted epitopes and HLA-II restricted epitopes respectively. The results of evaluations confirmed that the proposed vaccine is a 60.3 kDa stable, water soluble and high antigenic protein with high affinity to the selected target molecules and could elicits both humoral and cellular responses.

Altogether, the study suggests that the proposed vaccine can be considered as an effective anti-COVID-19 vaccine candidate for Iranian population.

The ongoing global pandemic of coronavirus disease (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus- 2 (SARS CoV-2) has jeopardized our health system and leaving everyone in disarray.  Despite the diligent cumulative effort of academia, there is hardly any light in the end tunnel so far in developing efficient and sustainable treatment options to tackle this public health threat. Therefore, designing a suitable vaccine to overcome this hurdle calls for immediate attention. The current study aimed to design a multi-epitope based vaccine using immunoinformatics tools. We approached the structural proteins: S, E, and M proteins of SARS CoV-2 since they facilitate the infection of the virus into a host cell. By using different bioinformatics tools and servers, the multiple B-cell and T-cell epitopes were predicted potential for the required vaccine design. The phylogenetic analysis provides in-depth knowledge on ancestral molecular changes and the molecular evolutionary relationship of S, E, and M proteins. Based on the antigenicity and surface accessibility of the spike (S), envelope (E), and membrane (M) proteins, eight epitopes were selected by various B cell and T cell epitope prediction tools.  Molecular docking was executed to interpret the binding interactions of these epitopes from where three potential epitopes WTAGAAAYY, YVYSRVKNL, and GTITVEELK were finalized with their noticeable higher binding affinity scores -9.1,-7.4, and -7.0  kcal/mol, respectively. It is noteworthy to mention that the targeted epitopes are believed to cover 91.09% of the population coverage worldwide. In sum, we identified the three most potential epitopes at length, which might be turned to our purpose of designing the peptide-based vaccine against SARS CoV-2.

Coronavirus disease 2019 (COVID-19) is undoubtedly the most challenging pandemic in the current century with more than 293,241 deaths worldwide since its emergence in late 2019 (updated May 13, 2020). COVID-19 is caused by a novel emerged coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Today, the world needs crucially to develop a prophylactic vaccine scheme for such emerged and emerging infectious pathogens.

In this study, we have targeted spike (S) glycoprotein, as an important surface antigen to identify its B- and T-cell immunodominant regions. We have conducted a multi-method B-cell epitope (BCE) prediction approach using different predictor algorithms to discover the most potential BCEs. Besides, we sought among a pool of MHC class I and II-associated peptide binders provided by the IEDB server through the strict cut-off values. To design a broad-coverage vaccine, we carried out a population coverage analysis for a set of candidate T-cell epitopes and based on the HLA allele frequency in the top most-affected countries by COVID-19 (update 02 April 2020).

The final determined B- and T-cell epitopes were mapped on the S glycoprotein sequence, and three potential hub regions covering the largest number of overlapping epitopes were identified for the vaccine designing (I531–N711; T717–C877; and V883–E973). Here, we have designed two domain-based constructs to be produced and delivered through the recombinant protein- and gene-based approaches, including (i) an adjuvanted domain-based protein vaccine construct (DPVC), and (ii) a self-amplifying mRNA vaccine (SAMV) construct. The safety, stability, and immunogenicity of the DPVC were validated using the integrated sequential (i.e. allergenicity, autoimmunity, and physicochemical features) and structural (i.e. molecular docking between the vaccine and human Toll-like receptors (TLRs) 4 and 5) analysis. The stability of the docked complexes was evaluated using the molecular dynamics (MD) simulations.

These rigorous in silico validations supported the potential of the DPVC and SAMV to promote both innate and specific immune responses in preclinical studies.

New generation of allergy vaccines is capable of promoting the development of protective IgG and blocking the functionality of allergen-specific IgE. We incorporated universal and powerful T-cell epitopes from tetanus and diphtheria toxoids (TD epitope) into recombinant Che a 2, the well-known allergic profilin of Chenopodium album, to determine its immunological properties.

The sequence and accordingly the structure of the recombinant Che a 2 was altered to generate a hypoallergenic variant (rChe a 2.rs). Moreover, TD epitope was incorporated to produce a novel vaccine that was nominated as rChe a 2.rsT.D. The effect of treatment with these variants was evaluated on the generation of allergen-specific IgG class, as well as lymphocyte proliferation in mice. Moreover, IgE-binding characteristics of the allergic patients’ sera were determined by ELISA and proliferation and cytokine production was measured in T-cells.

ELISA and dot blot revealed strong reduction of the IgE-reactivity of human sera to the variants of Che a 2 as compared to the wild-type molecule. Furthermore, Che a 2.rs and Che a 2.rsT.D induced much lower levels of IL5 and IL13 secretion from allergic patients’ PBMCs in comparison to wild-type Che a 2 protein. In mice, rChe a 2.rsT.D induced high titers of Che a 2-specific IgG antibody capable of blocking IgE-binding to rChe a 2 and induced lymphocyte proliferation more potently than rChe a 2.rs.

Collectively, incorporation of T-cell epitopes of tetanus and diphtheria into hypoallergenic vaccines can dramatically enhance anti-allergic immune mechanisms, particularly in poor responders.

Hydatid disease is a ubiquitous parasitic zoonotic disease, which causes different medical, economic and serious public health problems in some parts of the world. The causal organism is a multi-stage parasite named Echinococcus granulosus whose life cycle is dependent on two types of mammalian hosts viz definitive and intermediate hosts. In this study, enolase, as a key functional enzyme in the metabolism of E. granulosus (EgEnolase), was targeted through a comprehensive in silico modeling analysis and designing a host-specific multi-epitope vaccine. Three-dimensional (3D) structure of enolase was modeled using MODELLER v9.18 software. The B-cell epitopes (BEs) were predicted based on the multi-method approach and via some authentic online predictors. ClusPro v2.0 server was used for docking-based T-helper epitope prediction. The 3D structure of the vaccine was modeled using the RaptorX server. The designed vaccine was evaluated for its immunogenicity, physicochemical properties, and allergenicity. The codon optimization of the vaccine sequence was performed based on the codon usage table of E. coli K12. Finally, the energy minimization and molecular docking were implemented for simulating the vaccine binding affinity to the TLR-2 and TLR-4 and the complex stability. The designed multi-epitope vaccine was found to induce anti-EgEnolase immunity which may have the potential to prevent the survival and proliferation of E. granulosus into the definitive host. Based on the results, this step-by-step immunoinformatics approach could be considered as a rational platform for designing vaccines against such multi-stage parasites. Furthermore, it is proposed that this multi-epitope vaccine is served as a promising preventive anti-echinococcosis agent.

Leptospirosis is a widespread zoonotic disease caused by Leptospira interrogans. The conventional vaccines have some major problems. Therefore, recombinant vaccines such as multiple-epitope vaccine are suggested. OmpL1 and lipL32 are the most important proteins of Leptospira interrogans bacteria that can be used in epitope prediction process to design a multiple-epitope vaccine. Hence, in this study, the most reliable and accurate online servers were applied to predict B cell and T cell epitopes, the secondary and tertiary structures, enzyme digestion, and antigenicity score of ompL1 and lipL32. The results showed that epitopes located at 103 - 122, 210 - 232, and 272 - 291 amino acid residues are the common epitopes between T cell (MHCI) and B cell. 288 - 308 amino acid residues were introduced as common epitopes to stimulate both T cell (MHCI and MHCII) and B cell of ompL1 protein. In the case of LipL32 protein, 80 - 96 amino acid residues are recommended for T cell epitopes and 63-81 amino acid residues for stimulation of both B and T cells. All the mentioned epitopes can be considered as linear epitopes in designing a recombinant vaccine based on chimeric epitopes. It appears that these epitopes can be applied to design recombinant multiple-epitope vaccines against leptospirosis.

Neuraminidase (NA) is one of the surface proteins of influenza A virus, which plays an important role in immunization against influenza infection and is recognized as an important therapeutic target. Genetic and antigenic changes and substitutions can influence the efficacy of vaccine and change viral sensitivity to NA inhibitors (NAIs). In this study, we performed phylogenetic and molecular analyses of NA changes in influenza A(H1N1)pdm09 virus, compared them with the corresponding vaccine strain, and examined drug resistance mutations in isolates from patients.
The complete sequence of NA genes from 34 pandemic H1N1 isolates (identified in 2009-2010, 2010-2011, and 2013) was determined and analyzed both genetically and antigenically. The phylogenetic tree was plotted relative to the corresponding vaccine strain, using MEGA6 software package, based on the maximum likelihood method and JTT matrix (bootstrap value of 1000).
The phylogenetic analysis of pandemic isolates showed 31 amino acid substitutions in NA genes, compared to the vaccine strain . Some of these substitutions (N248D, V241I, N369K, N44S, and N200S) were important in terms of phylogenetic relationship, while the rest (D103N, V106I, R130T, N200S, G201E, and G414R) influenced the antigenic indices of B-cell epitopes. The catalytic sites, framework sites, and N-glycosylation remained unchanged in the studied samples. Meanwhile, H275Y substitution, related to oseltamivir resistance, was detected in 3 isolates. The average nucleotide identity of NAs with the corresponding vaccine strain was 99.415%, 98.607%, and 98.075% in 2009-2010, 2010-2011, and 2012-2013, respectively.
In this study, we provided basic information on the genetic and antigenic changes of NA genes in influenza A(H1N1)pdm09 virus from patients in 3 different seasons in Tehran, Iran. Considering the viral NAI resistance and changes in NA gene sequences of the isolates in comparison with the vaccine strain, further studies should be performed to monitor genetic changes in Iran. Moreover, the efficacy of vaccines should be examined.

Surface antigens (SAGs) of Toxoplasma gondii are known candidates for diagnostic tests and vaccines. The present study argues about the main necessary properties for determination and prediction of T-cell agretopes and B-cell epitopes of surface antigens of Toxoplasma gondii.
Primary, secondary and tertiary structures of the proteins were analyzed by different methods. The three-dimensional structures were determined by use of ab initio method for prediction of discontinues epitopes. The agretopes and epitopes were predicted via several various web servers with different methods employed.
The results of in silico analyses showed that the regions 129-GAPAGRNNDGSSAPT-143 for protein p22, 234-SENPWQGNASSD-245 for protein p30 and 348-PGTEGESQAGT-358 for protein p43, have the highest immunogenic potential.
We reached to three antigenic epitopes for cloning and protein expression. In following the purified polypeptide will be applied for diagnosis of Toxoplasma gondii.

Bovine Rotavirus and Bovine Coronavirus are the most important causes of diarrhea in newborn calves and in some other species such as pigs and sheep. Rotavirus VP8 subunit is the major determinant of the viral infectivity and neutralization. Spike glycoprotein of coronavirus is responsible for induction of neutralizing antibody response.
In the present study, several prediction programs were used to predict B and Tcells epitopes, secondary and tertiary structures, antigenicity ability and enzymatic degradation sites. Finally, a chimeric antigen was designed using computational techniques. The chimeric VP8-S2 antigen was constructed. It was cloned and sub-cloned into pGH and pET32a() expression vector. The recombinant pET32a()-VP8-S2 vector was transferred into E.oli BL21CodonPlus (DE3) as expression host. The recombinant VP8-S2 protein was purified by Ni-NTA chromatography column.
The results of colony PCR, enzyme digestion and sequencing showed that the VP8-S2 chimeric antigen has been successfully cloned and sub-cloned into pGH and pET32a().The results showed that E.coli was able to express VP8-S2 protein appropriately. This protein was expressed by induction of IPTG at concentration of 1mM and it was confirmed by Ni–NTA column, dot-blotting analysis and SDS-PAGE electrophoresis.
The results of this study showed that E.coli can be used as an appropriate host to produce the recombinant VP8-S2 protein. This recombinant protein may be suitable to investigate to produce immunoglobulin, recombinant vaccine and diagnostic kit in future studies after it passes biological activity tests in vivo in animal model and or other suitable procedure.

Designing an effective vaccine against human immunodeficiency virus (HIV)-1 is a global health priority. Multi-epitope vaccines offer several potential advantages that may be promising in case of mutable divergent pathogens such as HIV-1. Herein, a multiepitopic recombinant protein containing various HIV-1 antigens was expressed in E. coli cells and its immunogenicity in combination with different adjuvants was initially evaluated in BALB/c mouse. HIVtop4 sequence spanning the junction of six amino acid fragments (Gag158-186, Pol150-190, ENV296-323, ENV577-610, Tat1-20 and Tat44-61) was designed based on immunoinformatic analysis to reduce the creation of junctional epitopes, improve the cleavage of proteasome and avoid the local accumulation of hydrophobic regions. Synthesized nucleotide sequence corresponding to HIVtop4 was cloned into pET23a plasmid. Expression of pET-HIVtop4 plasmid was induced in BL21 (DE3) E. coli cells by addition of 1 mM IPTG during 3 h culture and the protein was purified by Ni-NTA column chromatography and further confirmed against anti-His antibody in western-blotting. Groups of BALB/c mice (n=6) were immunized three times with 2 weeks interval, subcutaneously with 10 m g of candidate vaccine adjuvanted in Complete Freund’s adjuvant, Montanide ISA70 and Alum with suitable control groups. Two weeks after last immunization lymphocyte proliferation was measured with Brdu, IL-4 and IFN- g cytokines with ELISA, total antibody and IgG1, IgG2a isotypes with indirect ELISA methods. Results showed that Immunization with HIV-1 tat/pol/gag/env led to a significant increase in the proliferative responses of lymphocytes, IL-4 and IFN-γ cytokine production and humoral immune response in comparison with the control groups. In this study we concluded that Tat, Env, Pol, Gag with adjuvants (Montanide, Alum and CFA) has potentials as a candidate vaccine against the HIV-1 virus.

Human cytomegalovirus is one of the most common pathogenic viruses all over the world. In congenital infection leads to neurologic severe disorders and even death of fetus and in individuals with immunosuppression may also cause severe clinical symptoms. Multiple evidence indicate that among several strategies, epitope-based vaccine (EVs) that can induce both humoral and cellular immunity responses, are the most important and have numerous potential profits. In this study, we select the viral surface glycoprotein B and phosphoprotein 65 and 150 with the highest antigenic and immunogenic properties, that have the most important role in induce cellular and humoral immune responses. Bioinformatics tools, as a standard and developed approaches use for epitope mapping. Epitope discovery greatly accelerate by in silico prediction methods with in vitro and in vivo verification. Bioinformatics methods and epitopes identification algorithms were used in order to selection of cytomegalovirus immunodominant epitopes, detection of each epitope antigenicity and design chimeric gene construct. The chimeric protein showed high antigenicity in vaxiJen analysis. Also further immunoinformatic analyses in order to predict the discontinuous and continuous B and T cell epitopes and MHC binding peptides affinity were used. The study results show that protein structures were suitable. Therefore it can be expected that construct is proper subject for practical experiments and stimulus for humoral and cellular immune responses.