فهرست مطالب

Archives of Medical Laboratory Sciences - Volume:3 Issue:1, 2017
  • Volume:3 Issue:1, 2017
  • تاریخ انتشار: 1395/12/30
  • تعداد عناوین: 6
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  • Maryam Bidram, Farida Behzadian, Fatemeh Fotouhi, Maryam Fazeli Page 1
    Background
    The influenza virus hemagglutinin is the major surface protein of the influenza A virus which is composed of HA1 and HA2 subunits. HA1 has an important role in binding of virus to cells and designing neutralizing antibodies. Escherichia coli (E. coli) and Bacillus subtilis (B. subtilis) both are known as the most useful prokaryotic hosts to express recombinant proteins. The aim of this study was to clone and express recombinant HA1 protein in E. coli and B. subtilis bacteria.
    Materials And Methods
    HA1 gene was cloned into pET-28a vector and pHT43 shuttle vector and then, both transformed to E. coli. The recombinant plasmids were extracted and then transformed into the BL21 and WB600 as expressing hosts. After induction with isopropyl-β-d-thiogalactoside (IPTG), expressed recombinant protein was analyzed by SDS-PAGE. Finally, the expressed protein was confirmed by the Western blot.
    Results
    HA1 gene was cloned into pET-28a vector and pHT43 shuttle vector and then, both transformed to E. coli. The recombinant plasmids were extracted and then transformed into the BL21 and WB600 as expressing hosts. After induction with isopropyl-β-d-thiogalactoside (IPTG), expressed recombinant protein was analyzed by SDS-PAGE. Finally, the expressed protein was confirmed by the Western blot.
    Conclusion
    This study demonstrated a strategy for production and purification of recombinant protein in large scale to test as vaccine candidate against influenza and it’s potentially immunogenicity be assessed in animal models.
    Keywords: Influenza A Virus, H3N2 Subtype, Hemagglutinin
  • Ehsan Khalili, Mohammad Reza Khoshayand, Ebrahim Abbasi, Mostafa Lakzaee, Mohammad Reza Parvizi, Mohammad Ali Faramarzi, Mahdi Aminian, Farshad Nojoomi Page 7
    Background
    Botulinum toxin, the most potent biological toxin, has become a powerful therapeutic tool for a growing number of clinical applications. Molecular studies have identified a family of synaptic vesicle-associated membrane proteins (VAMPs, also known as synaptobrevins) which have been implicated in synaptic vesicle docking and fusion with plasma membrane proteins.
    Materials And Methods
    Using the synaptobrevin as a substrate for in vitro assay is the method to detect BoNT activity. We have been working on optimizations of bacterial expression conditions and media for high-level production of synaptobrevin peptide. Statistics-based experimental design was used to investigate the effect of medium components (E. coli strain, peptone, IPTG, yeast extract, ampicillin, and temperature) on synaptobrevin production by E. coli.
    Results
    A 24 fractional factorial design with center points revealed that IPTG and temperature were the most significant factors, whereas the other factors were not important within the levels tested. This purpose was followed by a central composite design to develop a response surface for medium optimization. The optimum medium composition for synaptobrevin production was found to be: IPTG 29 mM, peptone 10 g/L, yeast extract 5 g/L, temperature 23°C and ampicillin 100 mg/L. This medium was projected to produce, theoretically, 115 mg/L synaptobrevin.
    Conclusion
    The optimum medium composition synaptobrevin production was found to be: BL21 (E.coli strain), LB medium (peptone 10 g/L, Yeast 5 g/L), Ampicillin (100 mg/L), IPTG (0.29 mg/L) and temperature (23°C).
    Keywords: synaptobrevin, E. coli, experimental designs, central composite design, medium optimization
  • Seyyed Shamsadin Athari Page 15
    Aim of Study: Adenosine deaminase (ADA) specifications (EC3.5.4.4) is an enzyme is involved in purine metabolism that breaks adenosine and deoxy-adenosine and produce inosine and deoxy-inosine and ammonia. The highest levels are of adenosine deaminase activity in monocytes and lymphocytes. High serum ADA activity with increased serum levels of AST, ALT and immunoglobulins were reported in variety of diseases including hepatitis. We aimed to investigate the activity of total ADA in serum of patients with hepatitis B and were determined molecular weight ADA1 and ADA2 isozymes in serum and RBC of hepatitis patients.
    Methods
    We were defining experiments by electrophoresis on SDS-PAGE, for isozymes of ADA; ADA1 and ADA2 in serum and red blood cells. ADA1 molecular weight was estimated at about 35 KDa and ADA2 about 110 KDa. The total ADA activity was measured by the modified Ellis method in 37 patients with hepatitis B and 40 healthy controls in the age range (20-60 years).
    Results
    The normal values for serum ADA in humans have been studied by various workers and found in serum samples to be in the range of 0-15 U/L, whiles in our analysis total ADA (tADA) enzyme activity is in controls and patients with hepatitis B, respectively, 13.35 ± 1.62 and 27.05 ± 8.49. Our results indicated that tADA level was higher in patients with hepatitis B than those of corresponding controls (P
    Conclusion
    Therefore, the serum ADA level could be used as an index along with other parameters in follow up of patients with hepatitis B.
    Keywords: Adenosine deaminase (ADA), hepatitis B, SDS-PAGE electrophoresis
  • Shiva Asadianfam, Nasibeh Abdollahi Page 21
    Background
    Diabetes is a type of metabolic diseases whose common characteristic is the increase in blood sugar. The prevalence of diabetes worldwide has progressively reached epidemic proportions. Its complications are microvascular and macrovascular complications, gastrointestinal and genitourinary disorders, infection and glaucoma. This study was designed to examine the incidence of diabetes in case of gender and age differences, and to investigate its complications .to better understand related etiology.
    Materials And Methods
    This research was a cross-sectional study undertaken on 380 diabetic patients referred to Amir-al-Momenin Hospital in Maragheh. The data were collected using patient records and analysed using SPSS software. The frequency and frequency percent of the qualitative data and the mean of the quantitative data were reported.
    Results
    The analysis suggested that among 380 diabetic patients, 248 were female and 132 were male, and 95.3% of those surveyed were married. The mean age of the patients under investigation was 57.66. 303 patients (79.7%) of the participants had a history of drug use, and the drugs are mainly cardiovascular drugs (35.8%), metformin (33.9%), and glibenclamide (28.2%). Over 50% of the participants had a history of illness, particularly cardiovascular disease (48.2%), while the least number of the participant had seizures. The highest and lowest number of patients had the blood sugar range of 424-325 (84 patients, 22.1%) and the blood sugar range of 622-523 (17 patients, 4.5%) , respectively. Moreover, the largest number of subjects (65 patients, 17.1%) had the blood cholesterol in the range of 325-271. About 75 participants had too much LDL. Besides, no statistically significant relationship was found between diabetes and white blood cells . 172 subjects developed complications. 70 patients, 68 patients, 55 patients, 51 patients, 14 patients, 13 patients, and 5 patients had renal, gastrointestinal, cutaneous, respiratory, cardiovascular, ocular complications, and amputations, respectively. About 110 subjects had various habits such as special diet (n = 44), smoking (n = 39), addiction (n=26) and drinking (n=3). Among 11 subjects of gestational diabetes mellitus (GDM), most had a high body mass index (BMI), the protein in the urine of about 18 percent of the subjects was positive, and regarding glucose disposal in the urine, 53 subjects were , 64 subjects were , 42 subjects were , 1 subject was ﯯ. Seventy participants had ketone disposal, which indicates kidney disorder.
    Conclusion
    The results revealed a higher prevalence of disease among middle-aged and older people in the society. The mean of B.M.I is 27.28 showing overweight in our population. The results also indicated a higher prevalence of hyperglycemia compared to hypoglycemia that may be due to very close relationship between diabetes and cardiovascular disorders, high levels of blood sugar in most of the diabetic patients. In addition, in a large number of fat people or people with hyperlipidemia were diabetic. In order to decrease mortality, and to diminish direct and indirect costs of diabetes, it is required to take essential measures. Such measures can include effective approaches to inform people for importance of nutrition education programs in controlling blood sugar and fat in diabetic patients via mass media and different programs.
    Keywords: diabetes, prevalence, diabetes complications, SPSS
  • Lida Moghaddam-Banaem Page 28
    Gestational diabetes, a metabolic disturbance in pregnancy has many predisposing factors such as maternal age, history of gestational diabetes, familial history of diabetes, and obesity. There have been many studies performed all over the world to recognize more risk factors for gestational diabetes in order to better control it. Micronutrients such as iron and zinc status in the body are good examples in this regard. The studies have shown that despite the increased need for iron during pregnancy, if there is an iron overload in body, it may predispose the mother to gestational diabetes; but there has been no significant relationship observed between body zinc status and gestational diabetes occurrence. Thus, medical practitioners should consider checking the iron status of pregnant women before giving iron supplements to them.
    Keywords: Gestational Diabetes Mellitus, Iron, Zinc, Pregnancy
  • Samira Rashidian, Mohammad Derakhshan, Ehsan Aryan, Roghayeh Teimourpour, Aida Gholoobi, Zahra Meshkat Page 30
    Background
    Novel tuberculosis (TB) vaccines that aim to boost and/or replace Bacillus Calmette-Guerin (BCG) are currently in development. DNA vaccines can stimulate both humoral and cell-mediated immunity in different animal models of TB and is thought to be a promising strategy in the development of new vaccines against TB. The aim of this study was to design and construct a DNA vaccine encoding ag85a and tb10.4 fusion genes of Mycobacterium tuberculosis.
    Materials And Methods
    tb10.4 fragment was amplified by PCR and the product was digested with restriction enzymes. Next, it was cloned into the pcDNA3.1 plasmid. The ag85a gene and pcDNA3.1ﻖ.4 plasmid were digested by EcoRI and BamHI restriction enzymes. Constructed vector was sequenced. The molecular analysis was done using bioinformatics software. New chimeric vector containing ag85a-tb10.4 genes were purified. Expression of pcDNA3.1ﻖ.4-ag85a plasmid was confirmed in eukaryotic cells.
    Results
    Fragments of 297 bp for tb10.4 and 1017 bp for ag85a were observed in agarose gel electrophoresis. Alignment of ag85a-tb10.4 genome sequence with reference genes in GenBank showed exact identities that indicate correction of all cloning procedures. Transfection of eukaryotic cells with pcDNA3.1ﻖ.4-ag85a vector and existence of tb10.4-ag85a fusion gene were both confirmed with RT-PCR.
    Conclusion
    In this study, tb10.4 and ag85a genes were isolated from Mycobacterium tuberculosis H37Rv strain and cloned into pcDNA3.1. Also, the capability of constructed vector in producing fusion ag85a-tb10.4 protein was confirmed with RT-PCR. pcDNA3.1ﻖ.4-ag85a vector can be used for further studies in future.
    Keywords: Mycobacterium tuberculosis, ag85a, tb10.4, pcDNA3.1+, DNA vaccine