S. pombe cells were grown in YE5S liquid medium at 25°C for 24 h, diluted 1:100, and grown an additional 12 h at 25°C to maintain a population in exponential growth phase at a density between 0.2 and 0.4 OD595. Equal numbers of cells were serially diluted 10-fold four times, and 5-μl aliquots were spotted onto YE5S agar plates for growth at 25, 30, or 36°C for 36–72 h. Plates were placed on a backlight box (Hall Productions, San Luis Obispo, CA), and a digital camera (Kodak DC290; Rochester, NY) was used to acquire images by transmitted light. Images were loaded into ImageJ, and colony growth was measured by densitometry using the NIH Image Gel Analyzer plug-in. Percentage growth for each condition is shown relative to wild-type growth in Table 2 .
Dc290
Manufactured by Kodak
Sourced in United States, Germany
The DC290 is a digital camera device designed for professional use. It features a high-resolution image sensor, advanced image processing capabilities, and connectivity options for transferring and managing images. The core function of the DC290 is to capture, store, and enable the transfer of digital images.
Automatically generated - may contain errors
Lab products found in correlation
Alpha digidoc 1000, by Bio-Techne (2 mentions)
Du530 life science uv vis spectrophotometer, by Beckman Coulter (1 mentions)
Rneasy mini kit, by Qiagen (1 mentions)
Coy anaerobic chamber, by Coy Laboratory Products (1 mentions)
Axiovert 25c, by Zeiss (1 mentions)
Se260, by Hoefer (1 mentions)
1d image analysis software, by Kodak (1 mentions)
9 protocols using dc290
1
Growth Analysis of S. pombe Cells
2
Histological Analysis of Liver Lipid Accumulation
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For our histology analysis, liver samples were fixed in 4% paraformaldehyde (PFA) at 4°C, cryoprotected overnight in 30% sucrose solution, embedded in Tissue-Tek OCT embedding compound, and frozen on dry ice. Next, 8-μm frozen sections were rehydrated, and neutral lipid accumulation was detected by Oil Red-O staining. Sections were rinsed with 60% isopropanol and stained for 20 min with prepared Oil Red O solution (0.5% in isopropanol followed by dilution to 60% with distilled water and filtered). After two rinses in 60% isopropanol and distilled water, slides were counterstained with hematoxylin for 4 min, rinsed with water, and mounted. Digital images were taken with a Nikon SMZ 1000 microscope connected to a Kodak DC290 digital camera.
3
DNA Ladder Visualization in Cells
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For DNA ladder visualization, control and treated samples (2.5 × 106 cells) were processed as reported [28 (link)]. Cells treated with 100 µM etoposide for 24 h were used as positive DNA ladder occurrence [31 (link)]. Pictures were taken with a photographic digital camera Kodak DC290 (Rochester, NY, USA).
4
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.
5
Detecting Staphylococcus epidermidis Biofilm
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DNA was extracted by the boiling method of Pérez-Pérez and Hanson 18 . The 16S rRNA gene sequence was determined to verify the presence of CoNS bacterial DNA, and then the icaACD genes were detected by PCR, which is considered the gold standard for detecting biofilm formation ability 14, 19 . The primers used, which are shown in Table 1, were obtained from GBT Oligos® and were designed based on the Staphylococcus epidermidis icaACD sequence (GenBank accession number U43366.1). To amplify the icaACD genes, a Thermo Cycler (model 2720, Biosystems) was used and programmed with the following cycling conditions: an initial step at 94°C for 5 min followed by 50 cycles of 30 sec each at 94°C, 55.5°C, and 72°C, with a final step at 72°C for 1 min. Each PCR simplex reaction contained 17.05 µL of ultrapure water, 1.75 µL of buffer (10×; Ludwig-Biotec®), 0.75 µL of MgCl 2 (50 mM; Ludwig-Biotec®), 2 µL of dNTPs (100 mM; Ludwig-Biotec®) s 0.2 µL of Taq DNA polymerase (5U/mL; Ludwig-Biotec®), 0.625 µL of forward primer, 0.625 µL of reverse primer, and 2 µL of DNA. The DNA fragments were analyzed by 1.5% agarose gel electrophoresis, and the bands were visualized with a photo documentation system (KODAK DC 290, using 1D software, version 3.6).
6
Anaerobic Enzyme Activity Assay
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S. oneidensis wild type and mutant strains were grown anaerobically in basal medium supplemented with 50 mM lactate, 0.02% casamino acids and 10 mM sodium sulfite or 0.5 mM potassium nitrite as electron acceptors. Cell extracts were prepared using B-PER lysis reagent (PIERCE, Rockford, IL). Protein concentrations were determined using the Coomassie Plus Protein Assay kit and 50 μg total protein was separated on 10% native polyacrylamide gels. Enzyme activities were assayed essentially as previously described41 (link). Gels were transferred to a Coy anaerobic chamber (Coy Laboratory Products, Grass Lake, MI) and stained for 20 min with reduced methyl viologen [38.9 mM methyl viologen dichloride and 57.4 mM sodium hydrosulfite in 10 mM Tris pH 7]. 10 mM of Na2SO3 or KNO3 was added and the gels were incubated further until bands of clearing, indicating reduction of the electron acceptors, were observed. Gels were imaged using a Kodak DC290 digital imaging system (Eastman Kodak, Rochester, NY).
7
Leptin Effects on In Vitro Wound Healing
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In order to study the effects of leptin on wound fill, an established in vitro wound healing model was used, as described in our previous experiments [44 (link), 45 (link), 47 ]. Briefly, cells were grown until confluence and 3 mm wide wounds, that is, cell-free areas, were created in a standardized manner in the cell monolayers. The wounded monolayers were treated with leptin (3 ng/mL) in the presence and absence of EMD for 4 d. Every day, the wounds were documented by inverse microscopy (Axiovert 25 C, 5x objective, Carl Zeiss, Oberkochen, Germany) and digital photography (Kodak DC 290, Kodak, Stuttgart, Germany). Afterwards, measurement and analysis of the wound widths were performed with special software (Alpha DigiDoc 1000, Alpha Innotech, San Leandro, CA, USA).
8
Wound Healing Assay with Adiponectin
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As previously described, an established in vitro wound healing model was used to analyze the effects of adiponectin on the wound closure in PDL cell cultures [43 (link)–46 ]. Briefly, cells were grown until confluence and 3 mm wide wounds, that is, cell-free areas, were created in the cell monolayers. The wounded cell monolayers were then exposed to adiponectin (3 μg/mL) in the presence and absence of EMD and documented by inverse microscopy (Axiovert 25 C, 5x objective, Carl Zeiss, Oberkochen, Germany) and digital photography (Kodak DC 290, Kodak, Stuttgart, Germany) over 4 d. Afterwards, the wound widths were measured and the wound closure was determined with special software (Alpha DigiDoc 1000, Alpha Innotech, San Leandro, CA, USA).
9
Myofibrillar Protein Analysis by SDS-PAGE
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The analysis of myofibrillar protein was done by using SDS-PAGE (SE 260, Hoefer Pharmacia Biotech Inc., USA). Gel was consisted by Stacking gel 4%, separating gel 10%. The sample of myofibrillar protein was loaded 10 µg per lane, for running buffer, upper running buffer (0.1 M tris, 0.15 M glycine, 0.15% SDS) and lower running buffer (0.025 M tris, 0.2 M glycine, 0.1% SDS) were separately manufactured by modifying Laemmli (1970) (link) process and used.
The operating condition of electrophoresis running was fixed by 4℃ and 40 mA, and 2 h operated (Rivero et al., 1997 (link)). To dye gel 0.05% Coomassie blue R-250 (w/v), 40% methanol and 7% acetic acid we used for 2 h at room temperature, so that to achieve sufficient dying, and 2 times bleaching were made by 40% methanol and 7% acetic acid.
To take gel image photo of myofibrillar protein Kodak DC290 (Eastman Kodak Company, USA) was used, and to analyze gel image Kodak 1D Image Analysis Software (Eastman Kodak Company, USA) was used (Huff-Lonergan et al., 2002 ; Ryu et al., 2005 (link)).
The operating condition of electrophoresis running was fixed by 4℃ and 40 mA, and 2 h operated (Rivero et al., 1997 (link)). To dye gel 0.05% Coomassie blue R-250 (w/v), 40% methanol and 7% acetic acid we used for 2 h at room temperature, so that to achieve sufficient dying, and 2 times bleaching were made by 40% methanol and 7% acetic acid.
To take gel image photo of myofibrillar protein Kodak DC290 (Eastman Kodak Company, USA) was used, and to analyze gel image Kodak 1D Image Analysis Software (Eastman Kodak Company, USA) was used (Huff-Lonergan et al., 2002 ; Ryu et al., 2005 (link)).
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