The sample was transferred to a 250 ml conical flask kept in water bath for alkali treatment. 75 ml of 17.5% caustic soda was measured using a measuring cylinder at 20°C. 15 ml of 17.5% NaOH was added and fibres were macerated gently with a flattened glass rod for 1 minute. 10 ml more NaOH was added and the solution was mixed for 45 seconds. 10 ml NaOH was again added and mixed for 15 seconds to make lump free slurry.
2.2.Yeast cell preconditioning and inoculum preparation 1 g dry weight of yeast was resuspended in 100 mL of deionized water in an Erlenmeyer flask of 250 mL volume, at 30–35°C, for 30 min with NaCl 6% w/v. Inoculum for experimental fermentations was prepared
The DEAE CL-6b gel was washed twice with 0.5m Hcl, twice with 0.5m Naoh and twice with PB ph6.0 before utilization. For DEAE 1ml gel every 1ml serum was utilized. In the wake of blending for 1 hour at 200c the suspension was centrifuged at 4500g for 25 minutes. The supernatant was thought by Ammonium sulfate precipitation and pellet was broken up in refined water and the ensuing arrangement was dialyzed against PBS ph7.2 to evacuate Ammonium sulfate and conformed to the first volume with yhe same cushion (Hong et.al., 1994). 2)
The supernatant was assayed for SOD activity by following the inhibition of epinephrine auto-oxidation. 0.5ml of sample was diluted with 0.5 ml of distilled water, to this 0.25 ml ethanol, 0.5 ml of chloroform (all reagents chilled) was added. The mixture was shaken for 1 min and centrifuged at 2000 rpm for 20 min. The enzymatic activity in supernatant was determined. To 0.05 ml of carbonate buffer (0.05 M, pH 10.2) and 0.5 ml of EDTA (0.49 M) was added.
21 Fresh semen samples were diluted with phosphate-buffered saline (PBS) to 2x106 sperm/mL. Fifty µL were incubated with 100 µL lysing reagent for 15 seconds and then 2 mL of PI was added and mixed with tube. Tube acquisition was done by flowcytometry immediately after staining. The intensity of its emission corresponds to the DNA content. Flowcytometric analysis displays a constant and characteristic bimodal nonartifactual DNA pattern indicating the presence of two different populations.
H2SO4 • Catalyst: potassium sulphate (K2SO4), copper (II) sulphate pentahydrate (CuSO4.5H2O) • NaOH (40%) • 0.1 N HCl solution • Boric acid (4%) • Indicator: methyl red (200 mg make up to 100 ml with %95 ethanol ) 1 gram Feta Cheese was used as a sample in this experiment. First of all, the experiment was started with the digestion phase which was performed by mixing 10g potassium sulphate and 0.5 g CuSO4 solution that acts as catalyst, and added 20ml conc. H2SO4. The digestion unit temperature was set to 420±10°C and preheated for 15 minutes. Then, it was heated continuously at least 2 hours till a clear colorless solution was obtained.
Next, a basic stock solution was used to prepare various concentrations ranging from 1.0 x 10-8M to 1.0 x 10-1M by serial dilution. The tissue was washed by overflow with reservoir’s solution for 5 seconds to obtain baseline before adding 0.1ml, 0.3ml and 0.5ml for each concentration respectively into the tissue bath.The tissue’s peak response for each final bath concentration(FBC) was measured and recorded. Rmax and EC50 of histamine were recorded. Later, 5ml of 1 x 10-6 M of mepyramine was added into the reservoir containing 1000ml of Krebs-Henssleit solution to produce a FBC of 5.0 x 10-9M. It was equilibrated with tissue for 10 minutes by flushing into the organ bath.
The reaction was started by adding 0.1 ml of the crude extract to 0.5 ml of the reaction mixture, which contained 7 mm MgCl2, 5 mm Na2MoO4, 2mm Na2-ATP and 0.032 U ml-1 of sulphate-free inorganic pyrophosphate in 80 mm Tris–HCl buffer (pH 8.0). Another aliquot from the same extract was added to the
MATERIALS AND METHODS Bacterial strains and culture conditions Two S. aureus strains were used in the present study; S. aureus 8325-4 (SigB-) and SH1000 representing a SigB+.strain. Overnight cultures were grown in Luria Broth (LB) at 37°Cwith shaking at 150 rpm. Exposure to antibiotics was carried out as detailed below. Antibiotics Ciprofloxacin were purchased from Sigma-Aldrich CO. 10 mg/ml stock solution of antibiotic were prepared freshly with 0.1N HCl and stored at -20°C. During the experiment we diluted with sterile water 1:10 and 1:100 depending on the different drug concentration.
This solution was diluted with diluents to gae a concentration of 0.1 mg/ml solution each of Amoxicillin trihydrate. The HPLC method was applied to the solutions and the results obtained were shown in table 4.6.11. System suitability solution: 25.0 µg/mL each of of USP Amoxicillin RS in Diluent. Precision
After adding 20 microliters of proteinase K to the 1.5 ml tube, the sample was vortexed for 30 seconds to disrupt the pellet. The sample was then incubated at 56°C to lyse the tissue. The sample was checked every fifteen minutes and vortexed between each checking for an hour and a half until the tissue was completely lysed. The tissue sample was then again vortexed. Next 200 microliters of buffer AL was added and
By matching the 1st and 2nd position of the template by di-base probes, fluorescence was detected and the extra tail with fluorescent probe is cleaved out. After multiple cycles of ligation, detection and tail cleavages, the extended chain reached the end of the template. Then the whole extension chain is removed and a new starting primer switching down 1 nucleotide position binds onto the template for another cycle of reaction. Totally, five round of primer binding cycles are performed to complete the sequencing of each fragment. 3.