Overall, 40–80 embryos at 3 dpf were homogenized and dissociated into single cells with 0.25% Trypsin-EDTA (Invitrogen). Cells were counted. Comparable amount of cells (∼4 × 105 cells) from each group were used. Cells were rinsed twice with methionine-free and cysteine-free DMEM (Invitrogen) followed by incubation in the same medium at 28.5°C for one hour. S35-labeled methionine and cysteine (Perkin Elmer, Santa Clara, CA, USA) were added into the culture medium at 10 μCi/ml. After incubation for another 1 h, cells were collected by centrifuging at 4°C for 5 min. Cells were rinsed twice with ice-cold phosphate-buffered saline (PBS). The cells were resuspended in 100 μl lysis buffer containing 50 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1% Triton X-100, and proteinase inhibitor cocktail (Roche Diagnostics). Proteins in the lysate were precipitated by adding trichloroacetic acid (Sigma-Aldrich) and incubated for 30 min on ice. Proteins were recovered by vacuum filtration on cellulose filters (Whatman, Uppsala, Sweden) and dried out. The filters were dissolved with scintillation cocktail, and radioactivity from samples were measured with Liquid Scintillation Analyzer (Perkin Elmer). Meanwhile, protein concentration was measured using NanoDrop 1000 Spectrophotometer (Thermo Scientific).
Cellulose filter
Manufactured by Cytiva
Sourced in United Kingdom
Cellulose filters are porous membranes made from cellulose materials. They are used for filtration and separation processes in various laboratory applications.
Automatically generated - may contain errors
Lab products found in correlation
Whatman, by Cytiva (9 mentions)
Trypsin edta, by Thermo Fisher Scientific (1 mentions)
Trichloroacetic acid, by Merck Group (1 mentions)
Liquid scintillation analyzer, by PerkinElmer (1 mentions)
Nanodrop 1000 spectrophotometer, by Thermo Fisher Scientific (1 mentions)
Proteinase inhibitor cocktail, by Roche (1 mentions)
Rotavapor r 200, by Büchi (1 mentions)
Centrifugal concentrator, by Eppendorf (1 mentions)
Ase 150, by Thermo Fisher Scientific (1 mentions)
9 protocols using cellulose filter
1
Quantifying Protein Synthesis in Zebrafish Embryos
2
Preparation of Spinach Methanol Extract
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The fresh leaves of spinach (S. oleracea L.) were harvested in the autumn–winter season in an agricultural field of the Puebla Province, México, and identified by a botanist in the herbarium of The National Polytechnic Institute (IPN, México City, Mexico). Voucher specimen number 4532 was deposited in the herbarium of the National School of Biological Sciences of IPN.
The leaves were dehydrated and finely ground. The dried leaves were macerated with methanol at room temperature, filtered through cellulose filters (Whatman®, Maidstone, UK), and dried using a rotatory vacuum evaporator (BUCHI Rotavapor R-200, Switzerland) at 45 °C to yield a green gum (95.0 g/kg dry leaves). The SME was stored in the dark at 4 °C until use. For all assays, the extract was reconstituted in distilled water [11 (link)].
The leaves were dehydrated and finely ground. The dried leaves were macerated with methanol at room temperature, filtered through cellulose filters (Whatman®, Maidstone, UK), and dried using a rotatory vacuum evaporator (BUCHI Rotavapor R-200, Switzerland) at 45 °C to yield a green gum (95.0 g/kg dry leaves). The SME was stored in the dark at 4 °C until use. For all assays, the extract was reconstituted in distilled water [11 (link)].
3
Cellulose Filter Preparation for Air Pollution Sampling
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Cellulose filters (Whatman 41) are recommended for analytical procedures and used in air pollution analysis as a paper tape for impregnation in sampling procedures. The filters were cut into 2.6-cm diameter circles. This diameter of the cellulose filter allows it to fit inside the tube cap. Each filter was impregnated using an 80-μL aliquot of triethanolamine solution. The triethanolamine impregnated filter is placed inside one of the sampler caps with the aid of tweezers. The cap is replaced on the sampler. The passive sampler (Fig. 2d ) is ready for use. It should be stored in a closed zip-lock plastic bag and stored in a refrigerator until use.
4
Extraction and Analysis of Organic Contaminants in Water Samples
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The water samples were thawed and once at room temperature, all samples were filtered through cellulose filters (0.7 μm, 90 mm, Whatman; Maidstone, UK). Three replicates of 250 mL (effluent) or 100 mL (influent) were processed according to a previously validated method in our research group (González-Gaya et al., 2021 (link)) (see details in section S2 in SI). Briefly, the samples were extracted with 500 mg solid-phase extraction (SPE) cartridges consisting of cation exchange (100 mg, ZT-WCX), anion exchange (100 mg, ZT-WAX) and reverse phase (300 mg, HRX) sorbents for effluent samples, and with 250 mg SPE cartridges containing half of the above-described amounts for influent samples. The cartridges were conditioned using 5 mL of MeOH:EtOAc and 5 mL of Milli-Q water. Subsequently, each sample aliquot was loaded and were left to dry under vacuum before analytes elution using 12 mL of a MeOH:EtOAc mixture (1:1) containing 2 % ammonia and 12 mL of a MeOH:EtOAc mixture with 1.7 % formic acid. Both extracts were combined, evaporated to dryness using a Turbovap (Zymark, Hopkinton, USA) under a gentle nitrogen stream and reconstituted in 250 μL of MeOH:Milli-Q water (1:1, v:v).
5
FTIR-ATR Analysis of Polymer Surface Groups
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The functional groups on the surface of the polymer were detected by means of Fourier transform infrared spectroscopy coupled with Attenuated Total Reflection (FTIR-ATR). For this, each sample was filtered onto 25 μm-pore size cellulose filters (Whatman), and 2-3 particles of each replicate were analysed. In order to remove possible bacterial aggregates, which could interfere in the FTIR-ATR analysis, the filters were previously cleaned with ethanol (70 % v/v) and left to dry in the stove (50 • C) for 15 min (Skariyachan et al., 2018) (link). MP particles were analysed in a Nicolet iN10MX micro-FTIR (Thermo Fisher Scientific; USA), by means of ATR, using a germanium tip. A Mercury Cadmium Telluride (MCT) detector cooled with liquid nitrogen was used. Spectra were collected in the middle infrared region (from 4000 to 675 cm -1 ), recording 16 scans at 16 cm -1 spectral resolution.
6
Determination of Metal Concentrations in Environmental Samples
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Samples of sediments, bivalve soft tissues, and Phragmites australis were subject to mineralization prior the analysis of metal concentration. 2.0 ± 0.1 g of sediment from each sample was placed in a conical flask and carefully flushed with 20 ml of HNO3 (14 mol L−1). Connected to a reflux condenser the flask was constantly heated at 80 °C for 30 min. After cooling the flask, its content was filtered using a cellulose filter (grade 1, 11 μm; Whatman, UK) into a polypropylene tube for analysis of metal concentration. A similar mineralization procedure was conducted for samples of Phragmites australis roots and leaves. Bivalves samples were kept for 14 days in containers flushed with HNO3 (14 mol L−1) then their contents were transferred to conical flasks and further prepared for metal concentration analysis as described for sediments and Phragmites australis samples. Procedure of sample digestion was adapted from EPA 3050B method (Environmental Protection Agency, USA).
7
Hydrogenation of Cbz-Protected Amine 80
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A round-bottom flask was charged with Cbz-protected
amine80 (175 mg, 0.33 mmol, 1 equiv), dry MeOH (3 mL),
and AcOH (0.3 mL). The flask was purged with N2 followed
by addition of Pd/C (10% w/w, 18 mg, 0.02 mmol, 5 mol %) and then
purging with H2 (balloon). The reaction was stirred for
2 days then filtered over a cellulose filter (Whatman), which was
washed with MeOH. The filtrate was concentrated under reduced pressure,
and the residue was purified using silica gel column chromatography
(2.5% → 5% MeOH/DCM with 5% Et3N) affording the
product (82 mg, 0.21 mmol, 63%). 1H NMR (400 MHz, CDCl3) δ 8.05 (t, J = 5.5 Hz, 1H), 7.35–7.26
(m, 2H), 7.26–7.17 (m, 3H), 6.74 (s, 1H), 3.84–3.76
(m, 2H), 3.75–3.58 (m, 4H), 3.30 (t, J = 6.5
Hz, 2H), 3.13 (s, 3H), 2.96–2.85 (m, 6H), 1.96 (s, 1H), 1.13–1.00
(m, 1H), 0.61–0.47 (m, 2H), 0.28 (q, J = 4.7
Hz, 2H). 13C NMR (101 MHz, CDCl3) δ 164.78,
163.74, 160.85, 156.51, 139.95, 128.80, 128.52, 126.20, 90.16, 51.66,
45.94, 45.26, 44.02, 35.62, 33.88, 10.84, 3.42. HRMS [C22H30N6O + H]+: 395.2554 calculated,
395.2558 found.
amine
and AcOH (0.3 mL). The flask was purged with N2 followed
by addition of Pd/C (10% w/w, 18 mg, 0.02 mmol, 5 mol %) and then
purging with H2 (balloon). The reaction was stirred for
2 days then filtered over a cellulose filter (Whatman), which was
washed with MeOH. The filtrate was concentrated under reduced pressure,
and the residue was purified using silica gel column chromatography
(2.5% → 5% MeOH/DCM with 5% Et3N) affording the
product (82 mg, 0.21 mmol, 63%). 1H NMR (400 MHz, CDCl3) δ 8.05 (t, J = 5.5 Hz, 1H), 7.35–7.26
(m, 2H), 7.26–7.17 (m, 3H), 6.74 (s, 1H), 3.84–3.76
(m, 2H), 3.75–3.58 (m, 4H), 3.30 (t, J = 6.5
Hz, 2H), 3.13 (s, 3H), 2.96–2.85 (m, 6H), 1.96 (s, 1H), 1.13–1.00
(m, 1H), 0.61–0.47 (m, 2H), 0.28 (q, J = 4.7
Hz, 2H). 13C NMR (101 MHz, CDCl3) δ 164.78,
163.74, 160.85, 156.51, 139.95, 128.80, 128.52, 126.20, 90.16, 51.66,
45.94, 45.26, 44.02, 35.62, 33.88, 10.84, 3.42. HRMS [C22H30N6O + H]+: 395.2554 calculated,
395.2558 found.
8
Accelerated Solvent Extraction of Malt Rootlets
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PLE was carried out using an accelerated solvent extractor system (ASE 150, Dionex, Sunnyvale, CA, USA). Sample was prepared by mixing 1.5 g of milled malt rootlets and 9 g of sand (used as dispersive agent) which was displayed in a stainless-steel extraction cell (10 mL). A cellulose filter (2.5 cm diameter, Whatman) was fixed at the cell bottom to avoid particles filtration. Initial equipment conditioning involved preheating at 1500 psi for 6 min followed by 100 s of nitrogen purge. Solvents were previously degassed in an ultrasonic bath for 30 min. Optimization of extraction conditions (extraction time, percentage of EtOH, and extraction temperature) was performed by a Box-Behnken incomplete factorial experimental design using the protein content as response variable and fitting the data to a quadratic equation similar to the previously shown for the optimization of UAE variables. As previously, the fitting of the regression model was assessed by ANOVA and the determination of R2.
All extracts were evaporated in a centrifugal concentrator (Eppendorf AG, Hamburg, Germany) and stored at −20 °C until use. Before use, solid samples were solubilized in a 5 mM phosphate buffer (pH 8).
All extracts were evaporated in a centrifugal concentrator (Eppendorf AG, Hamburg, Germany) and stored at −20 °C until use. Before use, solid samples were solubilized in a 5 mM phosphate buffer (pH 8).
9
Beetroot Pigment Extraction Protocol
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Fresh red beetroots were purchased from a local market. Unpeeled beetroots were finely grated and mixed with ethanol:water (4:1) for extraction of pigments, under continuous mechanical stirring for 1h in the dark. Typically 300 mL of solvent was used per 100 g of beetroot. The mixture was centrifuged at 30,000 g for 15 min and the supernatant was removed to be re-centrifuged once again. The new supernatant was then filtered using a cellulose filter (pore size 2 µm, Whatman). Ethanol was evaporated under reduced pressure and the resulting crude extract was filtered through an hydrophilic polypropylene membrane (pore size 0.2 μm, Pall Corporation). The crude extract was stored at -20 o C and diluted to the required concentration with Milli-Q water just prior use.
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