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13 protocols using fluorescent dna quantitation kit

1

ELISA Quantitation of Protein Levels

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Antibodies and standards were sourced from R & D Systems, and the ELISA was performed according to the manufacturer's instructions. Data were normalized to DNA content/well measured with a Fluorescent DNA Quantitation Kit (Bio-Rad).
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2

Soil Microbial Community Analysis by DGGE

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DNA was extracted from soil samples using PowerSoil DNA Isolation Kit (MoBio Laboratories, Inc. Carlsbad, CA, USA) according to manufacturer's instructions. Quantification of total DNA was achieved using a VersaFluor fluorimeter and a Fluorescent DNA Quantitation Kit (Bio-Rad, Milan, Italy). PCR was performed using primers FR1-GC (5′-(GC) AICCATTCAATCGGTAIT-3′) and FF 390 (5′-CGATAACGAACGAGACCT-3′) according to Mello et al. (2010 (link)).
Gels were stained with ethidium bromide solution (5 μg/mL; 20 min), washed with deionized water, and viewed by UV transillumination. Conversion, normalization, and further analysis of the DGGE patterns were carried out with Fingerprinting II Informatix™ software program (Bio-Rad). Similarities among profiles were calculated using the Unweighted Pair-Group Method with Average (UPGMA) algorithm.
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3

Palmitate Oxidation in Adipocytes

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Assessment was performed as reported previously (18 (link)). Briefly, differentiated adipocytes were starved in glucose-deprived DMEM for 1 h. Cells were then incubated at 37°C in DMEM containing 0.3 mM palmitate (9,10-[3H]palmitate, 12 μCi/ml), 2% BSA, 0.25 mM l-carnitine, and 3 mM glucose for 3 h. Palmitate oxidation was assessed by measuring 3H2O produced in the incubation medium. Data were normalized to DNA content/well measured with a Fluorescent DNA Quantitation Kit (Bio-Rad).
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4

Fluorescent DNA Quantitation Assay

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Cell proliferation assays were performed as previously described.49 (link) The DNA content of the cells was determined using a Fluorescent DNA Quantitation kit (Bio-Rad Laboratories, Hercules, CA). For each analysis, three replicate wells were used, and at least three independent experiments were performed.
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5

Soil DNA Extraction and Quantification

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The soil samples were transported back to the laboratory on ice, each manually homogenized, and sieved to remove large objects and debris. DNA was extracted from each homogenized soil sample (<250 mg) using the BIO 101 Fast DNA Spin Kit for Soil® and FastPrep®-24 System homogenizer (MP Bio, Solon, OH). The Fluorescent DNA Quantitation Kit (Bio-Rad, Hercules, CA) and Modulus™ Microplate Multimode Reader (Turner Biosystems, Sunnyvale, CA) were used to quantify the extracted DNA. Samples were diluted to a working stock of 20 ng/μL. Lastly, a 1% agarose yield gel was run to assess the integrity and quality of the extracted DNA.
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6

Quantifying Nasal dsDNA after Liposome Delivery

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Nasal wash samples were collected at 1 to 16 h after intranasal administration of liposomes (400 nmol/mouse) by flushing with 100 µL of cold PBS. After centrifugation to remove cells/debris, the amounts of dsDNA in the nasal wash samples were measured by a Fluorescent DNA Quantitation Kit (Bio-Rad Laboratories, Hercules, CA, USA) according to the manufacturer’s instructions using a Synergy HTX Multi-Mode Microplate Reader (BioTek Instruments, Inc.).
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7

Quantifying Cellular Triglycerides

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Total cellular triglyceride levels in RIPKO-1 and RIPKO-L cells were determined with the triglyceride colorimetric assay kit (Cayman Chemical) according to the manufacturer's instructions. Data was normalized to DNA content measured using fluorescent DNA quantitation kit (Bio-Rad Laboratories).
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8

Breast Cancer Cell Growth Assay

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MCF-7:5C cells were harvested after treatment with vehicle (0.1% ethanol), E2 (10−9 mol/liter, 1 nM), Dex (10−6 mol/liter, 1 µM), MPA (10−6 mol/liter, 1 µM), NETA (10−6 mol/liter, 1 µM), R5020 (10−6 mol/liter, 1 µM), RU486 (10−6 mol/liter, 1 µM), 4-OHT (10−6 mol/liter, 1 µM), or combinations, in triplicate, for specified time. Media and treatments were replaced every three days. DNA content was measured as using the Fluorescent DNA Quantitation Kit (Bio-Rad, Hercules, CA).
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9

Microsatellite PCR Fingerprinting of DNA from Musts and Wines

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Total DNA was extracted directly from musts and wines using the PowerSoil DNA Isolation Kit (MoBio Laboratories). Ten milliliter of each sample were centrifuged to collect cells. The DNA was then extracted according to manufacturer's protocol. Quantification of total DNA was achieved using a VersaFluor fluorimeter and a Fluorescent DNA Quantitation Kit (Bio-Rad, Milan Italy). DNA was used as a template for microsatellite PCR fingerprinting, as described by Vaudano and Garcia-Moruno (2008 (link)). PCR amplifications were performed in a thermocycler (MyCycler, Bio-Rad Laboatories, Milan, Italy) with the following PCR programme: 4 min of initial denaturation at 94°C, 28 cycles of 30 s at 94°C, 45 s at 56°C, 30 s at 72°C and, finally, 10 min at 72°C. The products were run on a 2.5% (w/v) agarose gel 1 × TAE buffer at 100 V for 80 min. Gels were stained with ethidium bromide. 1-kb plus DNA ladder (Life Technologies, Milan, Italy) was used as marker for the gel normalization.
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10

Chondrogenesis Induction and Characterization

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Chondro-pellet cultures (0.25 × 106 MSC) were induced towards chondrogenesis for 21 days with serum-free MesenCult-ACF differentiation medium (STEMCELL Technologies Inc, Vancouver, Canada). Harvested pellets were cryosectioned and 6-μm frozen sections stained with 1% toluidine blue (Sigma). Sulfated glycosaminoglycans (sGAG) were quantified after pellet digestion at 65 °C overnight with 1 mg/ml papain (Sigma) solution using the Blyscan Glycosaminoglycan Assay (Biocolor, Carrickfergus, UK) according to manufacturer’s instructions. DNA content was quantified using Fluorescent DNA Quantitation Kit (Bio-Rad Laboratories, Hercules, CA) according to manufacturer’s instructions.
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