Bacterial strains are listed in Table 1 . Unless otherwise stated, liquid cultures of Z. mobilis cells were grown semi-aerobically in Rich Medium (RM) [38 (link),39 (link)] without agitation at 30°C; in Falcon tubes (BD Biosciences), or Duran laboratory glass bottles (Schott AG), with caps that were fitted, but not air-tight; to allow limited gaseous exchange. Optical density measurements at 600 nm (OD600nm) were determined using a Beckman DU 530 Life Science UV/Vis spectrophotometer (Beckman Coulter Inc., USA). E. coli strains were grown aerobically in Luria Broth (LB) at 37°C. For agar plate preparation, 1.5% w/v agar was added. Plates were incubated aerobically at 30°C for Z. mobilis strains, or 37°C for E. coli strains. Antibiotics were used at the following concentrations: 100 μg/ml chloramphenicol (Cm) for Z. mobilis; 100 μg/ml ampicillin (Amp), 30 μg/ml Cm and 10 μg/ml tetracycline (Tc) for E. coli.
Du530 life science uv vis spectrophotometer
Manufactured by Beckman Coulter
Sourced in United States
The DU530 Life Science UV/Vis Spectrophotometer is a laboratory instrument designed to measure the absorption of ultraviolet and visible light by various samples. It is capable of analyzing the light absorption properties of a wide range of materials, including solutions, suspensions, and solid samples.
Automatically generated - may contain errors
Lab products found in correlation
Falcon tube, by BD (1 mentions)
Duran laboratory glass bottles, by Schott (1 mentions)
Tryptic soy broth (tsb), by Merck Group (1 mentions)
Yeast extract, by Thermo Fisher Scientific (1 mentions)
Autopure ls, by Qiagen (1 mentions)
Puregene dna isolation kit, by Qiagen (1 mentions)
Jsm 6700, by JEOL (1 mentions)
Jem 1010, by JEOL (1 mentions)
X pert mpd, by Philips (1 mentions)
Dc290, by Kodak (1 mentions)
Rneasy mini kit, by Qiagen (1 mentions)
Mpo assay kit, by Nanjing Jiancheng (1 mentions)
9 protocols using du530 life science uv vis spectrophotometer
1
Culturing Z. mobilis and E. coli Strains
2
Dual-species Bacterial Culture Protocol
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Cultures of E. faecalis and P. gingivalis were centrifuged and washed with phosphate-buffered solution (PBS) after the supernatant removal. Fresh Pg Broth (TSB, Sigma Aldrich; Yeast extract, Thermo Fisher Scientific™) was added to the pellet. Bacterial density was adjusted to 0.271–0.279 (2 × 108 CFU/mL) using a spectrophotometer (Beckman Coulter DU530 Life Science UV/Vis Spectrophotometer, Brea, CA, USA) at an optical density of 1 and a wavelength of 660 nm, according to MacFarland Standard scale no. 2. Then, 10-fold serial dilutions of the bacterial suspension were made up to 10−2 in tubes containing sterilized Pg broth. A total of 200 µL of the bacterial suspension was deposited into 96-well microtiter plates in quadruplicates, and 200 µL of sterilized Pg broth was deposited into the same well plates in triplicates to serve as the negative control. The same procedure was repeated in another two microtiter plates. For dual-species cultures, i.e., sequential or co-culture models, an equal volume of the two mono-species cultures were combined. All the specimens were incubated either aerobically or anaerobically for 96 h, and the culture medium was replenished after the first 48 h.
3
Tail Transection Assay for Hemostasis in Mice
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A 3 mm tail-tip transection method was used to assess haemostasis in 20–25 g male and female anaesthetized and ventilated mice22 (link). Following transection, the mouse tail was immediately immersed into warmed (37 °C) saline. The bleeding time was determined as the time from the tail transection to the moment the blood flow stopped for more than 120 s. A bleeding time beyond 30 min was considered as the cutoff time for the purpose of statistical analysis. Red blood cells were pelleted and lysed in 1 ml H20. Haemoglobin was quantified by absorbance at 575 nm (Beckman DU530 Life Science UV/Vis Spectrophotometer). Following cessation of bleeding, the incision was monitored for a further 120 s, and further bleeding within this period classified as ‘re-bleeding’. The duration of rebleeding was monitored as described above. Non-genotyped mice were used for the 3 mm tail-tip assay, and were subsequently genotyped using tail tissue collected post euthanasia.
4
Serotonin Transporter Genotyping Protocol
5
Biogenic Gold Nanoparticle Synthesis
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The purified fucoidanase (1 mL) was added to 10 mL of 1 mM aqueous solution of gold chloride and the solution was kept in a magnetic heater stirrer at 80 °C for 30 min, resulting in a color changing of solution that indicates the formation of gold nanoparticles. The formation of gold nanoparticles was monitored by UV-vis spectroscopy using Beckman Coulter DU530 Life Science UV/vis spectrophotometer (Beckman Coulter, Fullerton, CA, USA). Fourier transform infrared spectroscopy (FTIR) was performed by spectrum GX spectrometry in diffuse reflectance mode operated at a resolution of 4 cm−1 of wavelength of about 4000–400 cm−1. X-ray diffraction analysis (X’Pert-MPD, Philips, Amsterdam, The Netherlands) was performed by preparing a thin film of powdered gold nanoparticles. The studies on size, morphology and composition of the gold nanoparticles were performed by the means of field emission scanning electron microscopy (FESEM) (JSM-6700, JEOL, Tokyo, Japan) with energy dispersive X-ray analysis (EDXA) and high-resolution transmission electron microscopy (HRTEM) (JEM 1010, JEOL, Japan).
6
Measurement of Tail Bleeding Time
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Haemostasis was assessed, as described previously80 (link), by 3-mm tail amputation. The mouse tail was transected at 3 mm from the tip and immediately immersed into 37 °C saline. The bleeding time was determined as the time from the tail transection to the moment the blood flow stopped for more than 2 min. Bleeding time beyond 10 min was considered as the cut-off time for the purpose of statistical analysis. RBCs were pelleted and lysed in 1 ml H2O. Haemoglobin was quantified by absorbance at 570 or 575 nm (Beckman DU®530 Life Science UV/Vis Spectrophotometer). The investigator was blinded to the genotype during the experiment. Analysis of re-bleeding events: Following cessation of bleeding for 2 min, the incision was monitored for a further 2 min, and further bleeding within this period classified as ‘re-bleeding’. The duration of re-bleeding was monitored as described above. Haemostasis was measured in anesthetized mice (ketamine 100 mg/kg; xylazine 20 mg/kg I.P.).
7
Larval RNA Extraction and Analysis
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All microscopic analysis of spiracles and optical depressions was carried out using Leica Microsystems (Wetzlar GmbH, Wetzler, Germany).
7 Images were taken at 20 × 10.50 using a zoom digital camera (Kodak DC290, Tokyo, Japan).
Fifty fresh larvae were transferred to RNeasy lysis buffer that was supplied with the RNeasy® mini kit and the total RNA from larval samples was extracted using an RNeasy® mini kit (Qiagen Inc., Valencia, CA) and then measured with a Beckman DU® 530 Life Science UV/VIS spectrophotometer in accordance with the manufacturer's protocols. Sufficient amounts of RNA with adequate purity for subsequent analyses were obtained.
7 Images were taken at 20 × 10.50 using a zoom digital camera (Kodak DC290, Tokyo, Japan).
Fifty fresh larvae were transferred to RNeasy lysis buffer that was supplied with the RNeasy® mini kit and the total RNA from larval samples was extracted using an RNeasy® mini kit (Qiagen Inc., Valencia, CA) and then measured with a Beckman DU® 530 Life Science UV/VIS spectrophotometer in accordance with the manufacturer's protocols. Sufficient amounts of RNA with adequate purity for subsequent analyses were obtained.
8
Myeloperoxidase Activity Assay in Colon Samples
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MPO is an enzyme found mainly in granulocytes, and its activity was assessed using an MPO assay kit (Jiancheng Bioengineering, Nanjing, China) according to the manufacturer's instructions. Briefly, the MPO activities of the colon samples were detected by utilizing a kinetic assay in which H2O2 was degraded by the MPO released from neutrophil granulocytes. The absorbance at 460 nm was then measured using a DU 530 Life Science UV/Vis Spectrophotometer (Beckman Coulter, USA). The activity of the MPO is shown in units per gram of tissue (U/g tissue).
9
Quantifying Pseudomonas aeruginosa Pigment
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Overnight culture of Pseudomonas aeruginosa BIOTECH 1335grown in brain heart infusion broth (BHIB) was diluted to 0.06 OD at 600 nm using a UV-visible spectrophotometer (Beckman CoulterTM, DU®530 Life Science UV/Vis Spectrophotometer). 16 (link) Then, the 4.5 mL of P. aeruginosa culture was supplemented with 0.5 ml leave extract followed by incubation at 37°C for 24 hours. 16 (link) The treated culture was added with chloroform with the amount of 3 ml then mixing of the chloroform layer with 0.2 M HCl with a quantity of 1 ml. The extracted organic layer absorption was quantified with the used UV-vis spectrophotometer at 520 nm. For the blanks sample, sterile BHIB was used whereas P. aeruginosa culture of 4.5 ml with additional sterile distilled water of 0.5 ml was used as the control.
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