ABSTRACT
Staphylococcus aureus [S. aureus] is a major nosocomial pathogen and the infection with this organism in human is increasing due to the spread of antibiotic resistant strains. One of the resistance mechanisms of 5. aureus comprises modification in binding proteins to penicillin. Vaccine strategy may be useful in controlling the infections induced by this organism. This study aimed at developing and producing the recombinant protein PBP2a as a vaccine candidate and evaluating the related humoral immune response in a murine model. A 242 bp fragment of mecA gene was amplified by PCR from S. aureus COL strain and then cloned into prokaryotic expression vector pET-24a. For expression of recombinant protein, pET24alpha-mec cplasmid was transformed into competent E.coll BL2l [DE3] cells. Recombinant protein was over expressed with 1 mM isopropythio-beta-D-galctoside [IPTG] and purified using Ni-NTA agarose. SDS-PAGE and western blotting were carried out to confirm protein expression. For immunization of experimental groups, Balb/c mice were injected subcutaneously with 20 pg of recombinant PBP2a three times with three weeks intervals. The sera of experimental groups were collected three weeks after the last immunization and then specific antibodies were evaluated by ELISA method. Successful cloning of mecA was confirmed by colony-PCR, enzymatic digestion, and sequencing. SDS-PAGE and western blot analysis showed that recombinant protein with molecular weight of 13 kDa is over expressed. In addition, high titer of specific antibody against PBP2a in vaccinated mice was developed as compared with the control group and confirmed the immunogenicity of the vaccine candidate. Results suggest that PBP2a recombinant induced specific antibodies and can be used as Staphylococcal vaccine candidate after further studies
ABSTRACT
Iodine and Iodophore have been longley used as antiseptic to prevent infections with good efficacy and safety. The aim of this study was to formulate new povidone iodine 7.5% antiseptic foam and evaluate its antibacterial effect. In this experimental study, in the first step, the effects of the foaming agents including Sodium Lauryl Ether Sulfate [SLES], Sodium Lauryl Sulfate [SLS] and Nonoxynol 9 were studied and then the effect of pH and different buffer formulations including citrate, phosphate citrate and phosphate buffers on the organoleptic properties and the amount of the free iodine were evaluated according to the USP 32 criteria. The effects of the emollient such as glycerin and glycerox HE were evaluated and the accelerated stability test was taken for optimum formulations. Finally, the antimicrobial effect of the proper formulation was evaluated. SLES 3% in Citrate buffer with the pH 3.5 showed the best results for the quality of the foam and glycerin 1% was selected as proper emollient agent in presence of poloxamer 407 as foam stabilizer. The stability test was done on the superior formula and it proved its antiseptic properties. The foam formulation contained povidone iodine 7.5%, SLES 3%, Poloxamer [407] 2%, citrate buffer and glycerin 1% has passed stability as well as and antimicrobial effects tests; therefore, it can be used as a new antiseptic product and an appropriate detergent to prevent spreading of infectious disease
Subject(s)
Anti-Infective Agents, Local , Anti-Infective Agents , Chemistry, PharmaceuticalABSTRACT
Measles has been a major cause of illness and death in children and vaccination against the disease is part of the WHO global immunization program. A suitable vaccine should create maximum immune response against the pathogen and must be safe for the user. Thus, after production, vaccines must be analyzed and controlled by the producer and confirm by relevant governmental organizations. The Food and Drug Control Lab [FDCL], Ministry of Health, is the secondary control center on potency of vaccines in Iran. In this study, we have set up the WHO and NIBSC methods in FDCL and compare these methods on determining the potency of measles vaccine. Measles vaccines were obtained from Razi Institute Iran. Nine dilutions of vaccine [10[-1] to 10[-5]] in 0.5 log interval were mixed with Vero cell suspension and seeded. In WHO method, the cells were incubated at 36°C for 10 days, during which the cells were checked for cytopatic changes everyday. To set up the assay, we tested the vaccine dilution with four different cell suspensions [2x10[5]-5x10[4]/well] and four different concentration of serum [2.5-10%]. Based on our results, in the assays, 5% serum and 1x10[5] cells were used. The potency assay was performed with six different vaccines produced in one batch and the mean potency for Measles was 10[4.32 +/- 0.24] CCID[50]/vial for a ten-dose vial. In NIBSC method following seeding of Vero cells, the medium was removed after 3 hours and overlay was added. Then the plates were incubated at 35°C for 10 days. After incubation period, the overlay was removed, the plaques were stained with methyl violet and counted. This assay was repeated three times and the mean of the results was 5.83 +/- 0.03 log[10] PFU/dose. In this study, we have set up the WHO and NIBSC methods and results indicated that the potency of the vaccine is in acceptable range in either method. Furthermore, the WHO method is simple and less time consuming compared to NIBSC which is complicated and requires more effort to produce reproducible results