Three commercially available cocktails with phages against E. coli were used, Ses and Intesti of ELIAVA Biopreparations Ltd. (Tbilisi, Georgia) and Pyobacteriophag of Microgen Ltd. (Perm, Perm Territory, Russia) [32 (link),33 (link)]. The plaque assay was performed using the E. coli ATCC 29522 reference strain to determine the phage stock titer. Briefly, phage cocktail tenfold dilutions in 0.9% NaCl were made. Then, 50 μL of diluted phage stock were mixed with 100 μL of freshly grown bacterial suspension in 0.7% molten LB agar, previously incubated in LB broth for 16–18 h at 37 °C, and were poured onto a TSA plate. After overnight incubation at 37 °C, phage plaques on each plate were counted, and the phage titer (PFU/mL) was calculated. To achieve a higher phage cocktail titer, webbed plates of phage titration were used, and 12 mL of LB broth medium was poured on the plate and left at room temperature for 2 h. The supernatant with LB broth and overlay agar was removed and collected in 15 mL plastic tubes. Chloroform (CHCL3) was added with the final concentration of 2–3%, and the tubes were mixed and left at 4 °C for 2 h. Subsequently, the tubes were centrifuged at 6000× g for 15 min at 4 °C, and the supernatant was filtered using a 0.2 μm filter (Syringe filter, Filtropur S, Sarstedt, Nümbrecht, Germany).
Filtropur s
Manufactured by Sarstedt
Sourced in Germany
Filtropur S is a laboratory filtration device. It is designed for the filtration of liquids.
Automatically generated - may contain errors
Lab products found in correlation
Zetasizer nano z, by Malvern Panalytical (2 mentions)
Syringe filter, by Sarstedt (1 mentions)
Polypropylene tube, by Sarstedt (1 mentions)
Dmem high glucose, by Merck Group (1 mentions)
Nah13co3, by Merck Group (1 mentions)
Lck 314, by HACH (1 mentions)
Lck 514, by HACH (1 mentions)
Sulfuric acid, by Merck Group (1 mentions)
1200 series hplc system, by Agilent Technologies (1 mentions)
Varioskan lux spectrophotometer, by Thermo Fisher Scientific (1 mentions)
Zentrifuge z 36 hk centrifuge, by Hermle (1 mentions)
Genejuice transfection reagent, by Merck Group (1 mentions)
Pd miditrap g 25, by GE Healthcare (1 mentions)
Pd miditrap column, by GE Healthcare (1 mentions)
Microfuge 22r centrifuge, by Beckman Coulter (1 mentions)
Mini beadbeater, by Biospec (1 mentions)
Turbovap classic lv, by Biotage (1 mentions)
Vario el elemental analyzer, by Elementar (1 mentions)
Dx 500 dionex system, by Thermo Fisher Scientific (1 mentions)
19 protocols using filtropur s
1
Phage Cocktail Titer Optimization
2
Environmental Isolation of Staphylococcus hyicus
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Five different environmental samples were collected on the farm in parallel with sampling for S. hyicus. Two of these samples were liquid pig manure originating from two different pig houses on the corresponding farm. Another sample originated from washing water of a heavily infected piglet and a further sample originated from wastewater of a slatted floor. The last sample was a combination of washing water of an infected piglet and wastewater of a slatted floor. Two hundred and fifty milliliter of all five environmental samples were first centrifuged (14,300 × g, 15 min, 4°C) and then filtered through a 0.45 μm membrane filter (Filtropur S, Sarstedt, Nürnbrecht, Germany). A 5 ml aliquot filtrate of each of the five environmental samples was mixed with 100 μl CaCl2 (1 M) and 100 μl of a fresh overnight culture of the host bacterium S. hyicus (described in Isolation and Selection of Staphylococcus hyicus & Bacteriological Investigations). The mixture was incubated overnight at 37°C, then centrifuged at 18,000 × g for 15 min to remove residual bacterial cells. The supernatant was filtered again using a 0.45 μm membrane filter.
3
Quantitative Analysis of LAB
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The medium containing the cultured LAB was vortexed (MS1, IKA, Taquara, Brazil) until the pellet had dissolved, then centrifuged for 10 minutes at 5100 g in a Sigma 2–5 centrifuge and the supernatants were passed through a 0·20‐µm filter (Filtropur S, Sarstedt, Nümbrecht, Germany). The resulting liquid was injected directly into the HPLC without any previous dilution. Analyses were performed in triplicate for each bacterium.
4
Maize Root Exudates Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Seven-day-old maize seedlings in the paper roll system were transferred to the hydroponic system for another three weeks of cultivation. For exudate collection, four individual plants for each genotype (787, LH93, B73, lrt1 and rum1) were pooled and rinsed thoroughly three times with sterilized distilled water. Pooled roots were then immersed into a cylinder filled with freshly sterilized water with continuous aeration for 2-h collection. Root exudates were passed through a 0.2 µm filter (Filtropur S, Sarstedt) and frozen at -20°C. The root fresh weights were recorded in parallel for normalization of the exudates. The identification of apigenin and luteolin was performed as described above. In brief, 10 ml root exudates were first lyophilized and then resuspended in 250 µl of 50% MeOH containing 0.1% formic acid. Samples were sonicated for 15 min and shaken 1 h in an overhead shaker at 4 °C. After 30 min centrifugation (15,000 rpm, 4°C), 50 µl of the supernatants were subjected to UPLC analysis and measured values were normalized to root fresh weight and root exudate collection period and thus expressed as rate.
5
Gastrointestinal Digestion Stability Assessment
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The gastrointestinal digestion stability of extract in nonencapsulated and encapsulated forms was assessed (n = 6) according to a modified INFOGEST method [63 (link)]. For the gastric phase, pepsin was added (1.5 mL 34 mg pepsin/mL 0.1 N HCl–final conc. 5 mg/mL) to 2.5 mg of each phenolic compound (final volume of 8.5 mL), the pH was adjusted to 2, and the digests were incubated in the dark at 37 °C for 1 h (shaking at 120 rpm). For the small intestinal phase, final concentrations of 0.6 mg/mL lipase, 1.2 mg/mL pancreatin, and 3.6 mg/mL bile extract were added (prepared in 100 mM NaHCO3) to a final volume of 15 mL. The pH was adjusted to 6.5 using 1 N NaOH, and the digest overlaid with nitrogen gas and incubated, as described above, for 2 h. The final digests were centrifuged (4700× g, 60 min, 4 °C) and filtered through 0.2 μm membrane filter (Filtropur S, Sarstedt, Germany) to separate the bioaccessible fraction, respectively. Bioaccessible fraction was overlaid with nitrogen gas and stored at −80 °C before analysis. The total (poly)phenols or total antioxidant capacity was measured in the bioaccessible fraction and is displayed as a percentage of the total (poly)phenol and total antioxidant capacity in the original powders.
6
Ergot Extract Metabolism by MTHt3
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A suspension of MTHt3 was made by inoculating the strain in TSB and cultivating for 20 h, collecting biomass from 5 ml culture by centrifugation (2 655 × g; 10 min), and resuspending it in 5 ml 50 mM sodium phosphate buffer pH 7.0. Ergot extract was prepared by shaking 20 g ergot meal, obtained as described above, with 80 ml acetonitrile/water (1/1, v/v) for 2 h at room temperature. This crude extract was filtered (“White Ribbon Filter”, Grade 589/2: 4–12 μm; Whatman plc, Kent, UK), concentrated almost to dryness by rotary evaporation (Heidolph VV2011, Schwabach, Germany) and lyophilised (Christ, Osterode am Harz, Germany). Lyophilisate was dissolved (125 mg in 4 ml acetonitrile/water (1:1, v/v)) and filter-sterilised (Filtropur S, 0.2 μm, Sarstedt, Nümbrecht, Germany). One ml of this ergot extract was added to 0.08 ml MTHt3 suspension and 3.92 ml 50 mM sodium phosphate buffer pH 7.0, and the spiked suspension was incubated at 25°C and 200 rpm. Samples were taken and analysed by liquid chromatography/electrospray ionisation tandem mass spectrometry (LC/ESI-MS/MS) as previously described [37 (link)]. Negative control samples were taken from parallel incubation of the same, sterile ergot extract in the same buffer volume without MTHt3.
7
Microcapsule Preparation for In Vitro Testing
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Microcapsule suspensions for in vitro testing were prepared by suspending microcapsules at concentrations of 500 and 250 µg/ml26 (link) in Dulbecco’s Modified Eagle Medium (DMEM high glucose, D6429, Sigma Aldrich) with 100 µg/mL streptomycin and 100 U/mL penicillin.
The extract of SrRan loaded microcapsules was prepared as follows: 20 mg of microcapsules were placed into a 15 ml polypropylene tube (Sarstedt), and suspended in 5 ml of DMEM culture medium with 100 µg/mL streptomycin and 100 U/mL penicillin. The DMEM culture medium with streptomycin/penicillin was used as a control. The tubes were then incubated at 37 °C in a humidified atmosphere of 5% CO2 for 24 h. Then, the medium was collected and filtered using sterile syringe filters (Filtropur S, 0.2 µm, Sarstedt).
Obtained extracts and microcapsule suspensions were used for in vitro viability assay.
The extract of SrRan loaded microcapsules was prepared as follows: 20 mg of microcapsules were placed into a 15 ml polypropylene tube (Sarstedt), and suspended in 5 ml of DMEM culture medium with 100 µg/mL streptomycin and 100 U/mL penicillin. The DMEM culture medium with streptomycin/penicillin was used as a control. The tubes were then incubated at 37 °C in a humidified atmosphere of 5% CO2 for 24 h. Then, the medium was collected and filtered using sterile syringe filters (Filtropur S, 0.2 µm, Sarstedt).
Obtained extracts and microcapsule suspensions were used for in vitro viability assay.
8
Stable Isotope Labeling of Skeletonema costatum
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For global 13 C-labeling of S. costatum we used autoclaved artificial seawater medium that was prepared without addition of NaHCO3. An aliquot of this medium was utilized to dissolve NaH 13 CO3 (98 atom %, Sigma-Aldrich, Munich, Germany). This solution, containing sufficient NaH 13 CO3 to reach a final concentration of 2.38 mM, was sterile filtered (0.2 µm pore size, Sarstedt Filtropur S) and transferred back to the medium bottle. Tissue culture flasks were filled to the neck in order to minimize the area for CO2 exchange with the atmosphere and were inoculated with < 1 % (v/v) of a stationary S. costatum culture. After growing to stationary phase, an aliquot was taken and transferred to fresh 13 C-enriched medium (< 1 % (v/v)). After two of these cycles a plateau in the degree of labeling (verified by mass spectrometry as described below) was reached and the cultures were used for further experiments.
9
Analysis of Soluble and Total COD, VFAs, and Sulfides
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Samples for sCOD and VFA were centrifuged at 4500 rpm for 15 min at sample temperature, and supernatants were filtered with 0.2 µm syringe filters Filtropur S, SARSTEDT, Germany. The filtered samples for sCOD, and VFA, as well as unfiltered samples for TCOD were subsequently preserved by adding 1 % v/v of sulfuric acid (4 mol/L H 2 SO 4 , Merck Millipore, Germany), and maintained at +4 °C until analysis. VFAs were analyzed using a 930 Compact IC Flex 1 with a Metrosep A Supp 7 -250/4.0 column coupled with a 887 Professional UV/VIS detector (Metrohm, Sweden), and 0.1 M H 2 SO 4 was used as suppressor and 3.6 mN Na 2 CO 3 as eluent. The determination of TCOD and sCOD was performed using digestion vials LCK 514 and LCK 314, respectively (Hach Lange, Germany). Total dissolved sulfide concentration (S 2-) was analyzed using a modified methylene blue method, "Sulfulla" (see Supplementary material). In addition, pH, temperature and salinity of the sample were recorded for estimating the H 2 S fraction according to Eaton et al. (1995) .
10
Serum Protein Depletion Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Serum samples were diluted 5-fold with sodium phosphate buffer (phosphate-buffered saline—PBS), centrifuged at 16,000 rpm for 1 min in a Zentrifuge Z 36 HK centrifuge (Hermle Labortechnik Gmbh, Wehingen, Germany) at 4 °C, and filtered through a standard filter Filtropur S (Sarstedt, Nümbrecht, Germany) with a diameter of 22 microns. The resulting supernatant was passed through a Multiple Affinity Removal Column Human 14 (4.6 × 100 mm), Agilent, Santa Clara, CA, USA) for affinity binding and removal of 14 major proteins: albumin, IgG, IgA, transferrin, haptoglobin, antitrypsin, fibrinogen, alpha2-macroglobulin, alpha1-acid glycoprotein, IgM, apolipoprotein AI, apolipoprotein AII, complement C3, and transthyretin using an Agilent 1200 series HPLC system.
The resulting samples were concentrated via ultrafiltration through 3 kDa Microcon® centrifuge ultrafilters (Millipore, Molsheim, France) at 14,000 rcf for 15 min at 20 °C. The protein concentration was measured based on the absorbance at 280/260 nm using a Varioskan LUX spectrophotometer (Thermo Scientific, Waltham, MA, USA) located at the core facility Medical Genomics at Tomsk National Research Center.
The resulting samples were concentrated via ultrafiltration through 3 kDa Microcon® centrifuge ultrafilters (Millipore, Molsheim, France) at 14,000 rcf for 15 min at 20 °C. The protein concentration was measured based on the absorbance at 280/260 nm using a Varioskan LUX spectrophotometer (Thermo Scientific, Waltham, MA, USA) located at the core facility Medical Genomics at Tomsk National Research Center.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!