Mean Of Optical Density Research Articles

Expression of SARS-CoV-2 Nucleocapsid (N) Recombinant Protein Using Escherichia coli System

One of the main antigen that can be used for serological testing is the nucleocapsid (N) which is the most abundant viral-derived protein in SARS-CoV-2 where this virus can cause COVID19 disease. The aim of this study was to develop the SARS-CoV-2 N recombinant protein using Escherichia coli expression system. A total of 1,089 nucleotides encoding 362 amino acids of SARS-CoV-2 N was cloned to pET-14b vector. The plasmid then expressed in E. coli BL21 (DE3) and induced with 1.0 mM IPTG (Isopropyl-β-d-1-thiogalactopyranoside). The cell was harvested using denaturation lysis buffer due to inclusion body formation of SARS-CoV-2 N protein. Dialysis processed and concentrated using PEG-6000 resulted 0.992 mg/ml protein yield. Analysis of SARS-CoV-2 N recombinant protein using SDS-PAGE technique showed approximately 37.0 kDa specific band target protein. Application of this SARS-CoV-2 N recombinant protein to vaccinated and non-vaccinated antibody serum samples using ELISA technique indicated the significant result of optical density mean at 0.603 and 0.135, respectively. This study revealed that the production of SARS-COV-2 N recombinant protein could be carried out in E. coli expression system under denatured conditions, therefore the methods are more effective in producing the protein as a basic material in immuno-diagnostic assay.

Von Willebrand Factor Multimer Patterns in People Living with Human Immunodeficiency Virus Infection.

Endothelial dysfunction contributes to hypercoagulability in people with HIV (PWH). A surrogate marker of this is elevated von Willebrand factor (VWF) which has been documented in PWH. This study compared VWF multimer patterns in PWH who were anti-retroviral therapy (ART) naive and immune reconstituted on ART. VWF multimer analysis was performed with a semi-quantitative electrophoresis assay on plasma from 79 PWH (39 ART-naive and 40 virally suppressed on ART) and 25 normal control samples. The total optical density mean VWF level was significantly increased in PWH and higher in ART-naive versus ART-exposed, virally suppressed patients (p < 0.0001). No quantitative difference was demonstrated in the multimer distribution pattern between the groups. Our findings suggest that there is no difference in multimer distribution between ART-naive and virally suppressed PWH. Due to a small sample size, confirmation in a larger study is required.

EVALUATION OF SOX2 AS A POTENTIAL STEM CELL MARKER IN BENIGN AND MALIGNANT ODONTOGENIC TUMORS

INTRODUCTION: Odontogenic tumors (OTs) constitute a wide range of lesions derived from the tooth forming apparatus and its remnants. These tumors are generally benign, yet some are aggressive and have locally invasive behavior with a high rate of recurrence and some are malignant. Tumor stem cells (TSCs) are a unique subpopulation of cells that possess the ability to initiate a neoplasm and sustain self-renewal. Epithelial stem cell (ESC) markers such as sex-determining region Y (SRY)-box 2 (SOX2) are capable of identifying these stem cells which are expressed during the early stages of tooth development. OBJECTIVES: To evaluate immunohistochemically the expression of the stem cell proliferating marker SOX2 in benign and malignant epithelial odontogenic tumors. MATERIAL AND METHODS: Immunohistochemical (IHC) study using the SOX2 antibody was done on 30 surgical specimens of benign and malignant epithelial OTs as study group and 5 normal tooth germ tissues as a control group. RESULTS: Malignant OTs biopsies showed intense immune-reactivity to SOX2, while aggressive benign OTs showed medium immunereactivity with variable intensity. Totally benign tumors were immune-negative. The IHC staining intensity was directly proportional to the state of malignancy of odontogenic tumors. The difference in the expression of SOX2 in malignant and benign OTs was statistically significant. However, the difference in the expression of SOX2 in OTs and control group was not statistically significant in mean of area percent, while it was statistically significant in mean of optical density. CONCLUSIONS: SOX2 is highly expressed in malignant OTs biopsies. Medium immune-reactivity with variable intensities is seen in aggressive benign OTs.

Hemostatic Effect of the Combination of Factor VIII Concentrate with Tranexamic Acid (TXA) in the Prophylactic Setting in Severe Hemophilia a

IntroductionHemophilia is an X linked disorder characterized by an increased tendency to spontaneous bleeding resulting from profound compromise of the hemostatic response with delayed development of a clot and the formation of clots that are vulnerable to fibrinolysis. Optimization of clot stability by adding TXA to low concentrations of factor concentrates may be a way to improve prophylaxis in patients with severe hemophilia (sH) without substantially increasing the cost or the burden of more frequent infusions.AimsWe proposed to study whether hemostasis is improved with the addition of TXA to low FVIII plasma concentrations (equivalent to trough levels in prophylactic setting). We specifically studied in vitro the clot stability in patients with sHA; and in vivo the hemostatic response to trauma-induced joint bleed and tail clip in FVIII knock-out (KO) mice.MethodsAfter obtaining informed consent, blood samples of 12 adults with sHA were spiked in vitro to achieve final concentrations (FC) of 0-3-10 and 30 IU/dL of FVIII, then studied with and without TXA at FC 0.1mg/ml. Whole blood (WB) was assessed with ROTEM performed after addition of tPA in 6 patients. Thrombin generation (TG) was measured in plasma (PL) of 6 other patients. Clots obtained from TG were examined under electron microscopy (EM) to evaluate the quality of fibrin clot structure. Diameters of randomly selected 100 fibrin fibers were measured in each clot.In animal studies, 13 FVIII KO mice were given FVIII to a plasma concentration of 3 IU/dL to mimic trough FVIII levels in prophylactic setting; 7 were also infused with TXA (FC 0.1mg/ml). 10 minutes after infusion, joint bleeding was induced by knee puncture using a standardized protocol. 2 days post trauma, MRI was done and the joint bleed (surface of hyper signal into the joint) was compared between the groups with and w/o TXA.10 other FVIII KO mice were infused with 3 IU/dL rFVIII, of which 5 were also infused with TXA (FC 0.1mg/ml). 10 minutes after infusion, the extremity of the tail was cut at 2mm, and then immersed in pre-heated water-filled tube at 37°C. Blood was collected during 10 minutes. After lysis of erythrocytes, the amount of hemoglobin in collected blood was measured using spectrophotometry; means of optical density (OD) were compared between the 2 groups.ResultsA dose dependent improvement of TG was observed after adding FVIII alone (p=0.024). As expected, the addition of TXA had no effect on TG capcity. Fibrin fiber diameters were significantly decreased with TXA+FVIII compared to FVIII alone (results shown in fig1), suggesting a stronger fibrin network, as thin fibrin fibers are characteristic of a robust fibrin mesh. Surprisingly, ROTEM was fully normalized after addition of TXA only, with no exogenous FVIII.In FVIII KO mice, the addition of TXA to 3 IU/dL rFVIII significantly decreased joint bleeding compared to 3 IU/dL rFVIII alone (p=0.022) (fig2).With the tail clip experiment, the addition of TXA to 3 IU/dLFVIII showed a trend toward improved bleeding when compared to 3 IU/dLFVIII alone; with means of OD±SEM 0,31±0,19 and 0,94±0,45 respectively, but did not reach statistical significance (p=0,15) (fig3).ConclusionOur in vitro and in vivo results suggest a potential benefit of TXA when used in combination with FVIII in prophylactic setting. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Relationship between the Expression of α-syn and Neuronal Apoptosis in Brain Cortex of Acute Alcoholism Rats

To observe the changes of expression of α-synuclein （α-syn） and neuronal apoptosis in brain cortex of acute alcoholism rats and to explore the mechanism of the damage caused by ethanol to the neurons. The model of acute alcoholism rat was established by 50% alcohol gavage. The α-syn and caspase-3 were detected by immunohistochemical staining and imaging analysis at 1 h, 3 h, 6 h and 12 h after acute alcoholism. The number of positive cell and mean of optical density were detected and the trend change was analyzed. The variance analysis and t-test were also performed. The number of α-syn positive cell and average optical density in brain cortex of acute alcoholism rat increased significantly and peaked at 6 hour with a following slight decrease at 12 h, but still higher than the groups at 1 h and 3 h. Within 12 hours after poisoning, the number of caspase-3 positive cell and average optical density in brain cortex of rats gradually increased. The abnormal aggregation of α-syn caused by brain edema and hypoxia may participate the early stage of neuronal apoptosis in brain cortex after acute alcoholism.

Deparaffinization of formalin-fixed paraffin-embedded tissue blocks using hot water instead of xylene

This study aimed to deparaffinize formalin-fixed paraffin-embedded (FFPE) tissues using hot water instead of xylene and measuring the quantity and quality of the extracted DNA from the respective tissues. To deparaffinize the tissue sections with hot water, small sections were exposed to 90 °C distilled sterile water. After 25 FFPE tissue samples were deparaffinized with the hot water method, DNA was then extracted. The mean of optical density and the ratio of absorbance of the DNA solution were 220.01 ± 36.1 ng/μl and 1.65 ± 0.1, respectively. Polymerase chain reaction (PCR) analysis of the toll-like receptor 4(TLR4) gene showed that the method can be used as a tool for different applications.

The expressions of aquaporins in chronic stress depression model rat hippocampus and gastrointestinal mucosa

Objective To study the mechanisms of depression by exploring expressional differences of AQPs in the tissues of chronic stress depression rats. Methods Depression model was replicated by unpredicted chronic stress. 20 rats were randomly separated into normal control group and model control group, AQP4 and AQP3, AQP8 expressions on hippocampus and gastrointestinal mucosa of rat model with depression were detected by immunochemical staining method. Results Means of optical density of AQP4 of normal group and model group hippocampus were 0.28±0.02, 0.22±0.06 respectively, and the difference between two groups was significant statistically(t value was 2.756, P 0.05). Two groups' means of optical density of AQP3 on small intestinal mucosa were 0.15±0.02, 0.17±0.02, and the difference was significant statistically(t value was 2.769, P<0.05). Two groups' means of AQP8 optical density in gastric mucosa were 0.15±0.01, 0.19±0.04; 0.16±0.01 and 0.21±0.04 in small intestinal mucosa; 0.16±0.01 and 0.22±0.04 in colonic mucosa, and the differences were significant statistically(t values were 3.139, 5.113, 4.534, P<0.05, P<0.01, P<0.01). Conclusion The expression of AQP4 on depression model rat hippocampus are lower than those of the normal group; and the expression of AQP3 on gastric mucosa and colonic mucosa are no change obviously, but it's up-regulation on small intestinal mucosa. The expressions of AQP8 on gastric mucosa and small intestinal mucosa and colonic mucosa are up-regulating. Key words: Aquaporins; Gastrointestinal mucosa; Hippocampus; Rats; Depression

Kinetics of dengue virus NS1 protein in dengue 4-confirmed adult patients

In this work, the presence of NS1 protein as a possible early marker of dengue infection was studied in serum samples from confirmed adult patients with a primary and secondary dengue 4 infection. A total of 209 serum samples collected from day 2 up to day 7 of fever onset from 71 patients were tested by Platelia NS1 antigen capture ELISA kit (BioRad, Marnes-la-Coquette, France), and the results were compared with those obtained by capture antidengue virus IgM (MAC)-ELISA and ELISA inhibition method tests. The 83.3% of primary cases and 96.4% of secondary cases were NS1 positive. The kinetics of NS1 protein showed the highest values in optical density mean ratio or in percentage of positives between days 2 and 4. The results obtained in this study show the utility of the NS1 protein as a virologic early marker of dengue infection. Prospective studies should be carried out to confirm its utility as a prognostic marker of severe illness.

Cardiocyte Nuclear Chromatin Density Correlates With Transplanted Heart Left Ventricular Mass

Introduction Cardiocyte hypertrophy is accompanied by polyploidy, seen as a decrease in chromatin density in the enlarged nucleus. Repeated biopsies of a transplanted heart offer the possibility of a dynamic evaluation of these phenomena. The aim of this work was an evaluation of cardiocyte nuclear chromatin density in transplanted hearts during long-term follow-up. Materials and Methods The material encompassed myocardial biopsy specimens taken during the first week, first month, and then on an annual basis up to 10 years after surgery. Only biopsy specimens with no rejection were considered (grade “0” International Society for Heart and Lung Transplantation [ISHLT] 122 biopsy specimens). The control group consisted of 7 donor heart specimens. We evaluated the optical density—mean gray level—of cardiomyocyte nuclear chromatin. We determined correlations of this index with the nuclear area, and with left ventricle ultrasound measurements, using correlation analysis. Results The chromatin mean gray level decreased with time, correlating positively with interventricular septum thickness, left ventricle posterior wall diameter, and left ventricular mass. Analysis of individual periods showed a significant positive correlation of the mean grey level with the cardiocyte nuclear surface in year 3, 4, and 9 after transplantation, thereby suggesting the occurrence of polyploidy at those times. The significant negative correlation of these values (1 week and 1 year) indicated normalization of early cardiocyte hypertrophy. Conclusions With the passage of time chromatin condenses, leading to pyknosis. The activity of cardiocyte chromatin correlated with left ventricular hypertrophy. Compensatory cardiomyocyte polyploidy is a periodical phenomenon.

Studying the efficacy of giving anthrax toxin as a vaccine intranasally and subcutaneously

This study was concerned on the production of Bacillus anthracistoxin and its use as a vaccine. Two groups of guinea pigs wereimmunized with the toxin which was given in a dose of 0.5 mlsubcutaneously to the 1st group while the animals of the 2nd group wereimmunized by instillation in the nose 4 drops into each naris. The twogroups were given booster dose 14 days after the 1st dose. The immunizedand control animals were bled by cardiac puncture 20 days after thebooster dose.In the present study, anthraxin was used as an antigen in EnzymeLinked Immuno Sorbant Assay (ELISA) to measure the optical density(OD) obtained from adminsteration of vaccine in two different routes.High mean of optical density 1.660 ± 0.07 nanometer was recorded tosubcutaneous group and the mean 1.181 ± 0.002 nanometer to theintranasal group.

Therapeutic effect of naloxone on radiation optic neurapathy: Based on rabbit models

Background Radiation therapy is widely used to treat tumor of brain; however, irradiation of radiation into eye tissues may easily cause ischemia and hypoxia in retina and optic nerve tissue so as to induce radiation optic neurapathy. Noloxone is a specific antagonist of opiate receptor, and it can change injured effect of β-endorphin. Objective To observe the axoplasmic transport of optic nerve at various phases after radiation injury so as to investigate the mechanism and regularity of optic nerve injury; meanwhile, to verify the therapeutic effects of naloxone on radiation optic neurapathy. Design Randomized controlled animal study. Settings Medical College of Qingdao University, Changzhi Medical College. Materials A total of 40 healthy adult New Zealand rabbits, weighing 2–2.5 kg, of either gender, were checked by using slit lamp and ophthalmoscop before radiation in order to exclude eye diseases. FCC-7000 vertical kilocurie 60Co therapeutic machine was made in Yantai, China; in addition, naloxone was provided by Beijing Sihuan Pharmaceutical Factory. Methods The experiment was carried out in the Animal Experimental Laboratory, Medical College of Qingdao University from January 2005 to December 2006. The experimental rabbits were randomly divided into radiation group ( n = 16), treatment group ( n = 16), blank control group ( n = 4) and injured control group ( n = 4). Except blank control group, rabbits in other three groups were irradiated in as the center of optic chiasma including the area of optic nerve by using 60Co therapeutic machine. Radioactive source was 85 cm away from head, and the size of operative field was 5 cm×5 cm. The radiation was performed once a day with the dosage of 3 Gy for 8 days in total. The total dosage was 24 Gy. When radioactive dosage reached 24 Gy, 2 mg/kg naloxone was dropped into ear vein of rabbits in the treatment group once a day for 10 days in total. Rabbits in the injured control group were only irradiated but not given any drugs. Main outcome measures At 1, 10, 30 and 60 days after 24-Gy radiation, anterogradely labeled horseradish peroxidase (HRP) was used to measure optical density mean (OPTDM) in the radiation group and the treatment group; while, OPTDM was directly measured in the blank control group, and OPTDM was directly measured after radiation in the injured control group. Results All 40 experimental rabbits were involved in the final analysis. There were significant differences in OPTDM at various phases between radiation group and blank control group ( P < 0.05); in addition, there was significant difference in OPTDM in the treatment group at 1, 10, 30 and 60 days after 24-Gy radiation ( P < 0.05); otherwise, at 30 and 60 days after 24-Gy radiation, there was significant difference in OPTDM between radiation group and treatment group ( P < 0.05). Conclusion Naloxone may improve optic nerve axoplasmic transport disorder induced by radiation so as to protect optic nerve.

Transgenic potato plants expressing the potato virus X (PVX) coat protein gene developed resistance to the viral infection

The gene coding for potato virus X (PVX) coat protein (CP) was expressed in transgenic potato plants obtained byAgrobacterium tumefaciens transformation. One hundred independent clones were analyzed in challenge experiments for resistance to PVX infection under greenhouse conditions as a preliminary test. From this test, 16 clones with the best resistance results were selected for a small-scale field trial. Clones 54, 60, 73 and 91 demonstrated the best values of resistance to PVX in the field. Statistical analysis of the field trial showed significant differences between means of optical density obtained in ELISA from transgenic clones and non-transformed plants (P<0.05). There was correspondence between resistance to virus infection and expression of the CP gene of PVX virus in the analyzed clones.

Variations in Activity of the Human Natural Anti-Gal Antibody in Young and Elderly Populations

Anti-Gal is a natural antibody which constitutes approximately 1% of circulating IgG in humans and which interacts specifically with the mammalian carbohydrate epitope Gal alpha 1-3Gal beta 1-4GlcNAc-R (termed the alpha-galactosyl epitope). The purpose of this study was to determine whether there are age-associated changes in the affinity of anti-Gal in elderly individuals. The age-associated variations in affinity of anti-Gal were determined by the extent of this antibody binding to solid-phase alpha-galactosyl epitopes (mouse laminin) in ELISA, performed at various serum dilutions with serum samples from 140 elderly and 127 young individuals. The data were expressed as the mean of optical density (O.D.) values measured at serum dilutions of 1:40, 1:80, and 1:160, and correlated with the affinity of anti-Gal as measured in equilibrium dialysis. The proportion of individuals with low-affinity anti-Gal is sixfold higher in the elderly than in the young population. Whereas low-affinity binding of anti-Gal (mean O.D. value < or = 1.0) was observed in only 6% of the young population, as many as 35% of the elderly population displayed such low-affinity anti-Gal. Anti-Gal affinity in ELISA assay correlated with the affinity of this antibody as measured in equilibrium dialysis. The proportion of individuals with low affinity anti-Gal increases in the elderly population in comparison with the young population. The possible molecular mechanisms which may result in the age-associated decrease in the antibody affinity are discussed.