S. Typhimurium biofilms were grown on non-treated polystyrene 96-well plates (Corning, Kennebunkport, ME) by normalizing overnight (O/N) cultures grown in TSB to OD600 = 0.8, then diluting 1:10 into biofilm-growth media (TSB diluted 1:20), and dispensing 0.1 ml per well. The plates were incubated at 30 °C in a GyroMini nutating mixer (LabNet International, Inc., NJ) at 24 rpm. S. Typhi biofilms were grown as follows: O/N liquid cultures were incubated in TSB at 37 °C with aeration. These were then normalized to OD490 = 0.65, diluted 1:2500 in TSB, and 200 µL/well were dispensed into 8-well Permanox chamber slides (Thermo Fisher, Rochester NY) and incubated at 37 °C with 5% CO2. To simulate growth conditions on gallstones, wells were pre-coated with cholesterol by adding a solution of 5 mg/mL in 1:1 isopropanol:ethanol and air-dryed overnight17 (link). Media was changed daily for both S. Typhimurium and S. Typhi biofilm growth.
Gyromini nutating mixer
Manufactured by Labnet
Sourced in United States
The GyroMini nutating mixer is a compact laboratory device designed to provide gentle, orbital mixing motion. It is used to mix samples in tubes or microplates, ensuring thorough and consistent sample preparation.
Automatically generated - may contain errors
Lab products found in correlation
Non treated polystyrene 96 well plates, by Corning (2 mentions)
Spectramax spectrophotmeter, by Molecular Devices (2 mentions)
Permanox chamber slide, by Thermo Fisher Scientific (1 mentions)
Ph meter, by Sartorius (1 mentions)
Softmax pro, by Molecular Devices (1 mentions)
Screw cap tubes, by Avantor (1 mentions)
Softmax pro software, by Molecular Devices (1 mentions)
6 protocols using gyromini nutating mixer
1
Salmonella Biofilm Growth Protocols
Check if the same lab product or an alternative is used in the 5 most similar protocols
S. Typhimurium biofilms were grown on non-treated polystyrene 96-well plates (Corning, Kennebunkport, ME) by normalizing overnight (O/N) cultures grown in TSB to OD600 = 0.8, then diluting 1:10 into biofilm-growth media (TSB diluted 1:20), and dispensing 0.1 ml per well. The plates were incubated at 30 °C in a GyroMini nutating mixer (LabNet International, Inc., NJ) at 24 rpm. S. Typhi biofilms were grown as follows: O/N liquid cultures were incubated in TSB at 37 °C with aeration. These were then normalized to OD490 = 0.65, diluted 1:2500 in TSB, and 200 µL/well were dispensed into 8-well Permanox chamber slides (Thermo Fisher, Rochester NY) and incubated at 37 °C with 5% CO2. To simulate growth conditions on gallstones, wells were pre-coated with cholesterol by adding a solution of 5 mg/mL in 1:1 isopropanol:ethanol and air-dryed overnight17 (link). Media was changed daily for both S. Typhimurium and S. Typhi biofilm growth.
+ Expand
S. Typhimurium biofilms were grown on non-treated polystyrene 96-well plates (Corning, Kennebunkport, ME) by normalizing overnight (O/N) cultures grown in TSB to OD600 = 0.8, then diluting 1:10 into biofilm-growth media (TSB diluted 1:20), and dispensing 0.1 ml per well. The plates were incubated at 30 °C in a GyroMini nutating mixer (LabNet International, Inc., NJ) at 24 rpm. S. Typhi biofilms were grown as follows: O/N liquid cultures were incubated in TSB at 37 °C with aeration. These were then normalized to OD490 = 0.65, diluted 1:2500 in TSB, and 200 µL/well were dispensed into 8-well Permanox chamber slides (Thermo Fisher, Rochester NY) and incubated at 37 °C with 5% CO2. To simulate growth conditions on gallstones, wells were pre-coated with cholesterol by adding a solution of 5 mg/mL in 1:1 isopropanol:ethanol and air-dryed overnight17 (link). Media was changed daily for both S. Typhimurium and S. Typhi biofilm growth.
2
pH Measurement of Bacterial Growth Media
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The pH of the growth media was measured as follows. Before bacterial inoculation, the pH of 1:20 TSB with or without 0.2% or 2% L-arabinose was measured by a pH Meter (Denver Instrument). O/N cultures were normalized to OD600 = 0.8, diluted 1:10 into 20 mL 1:20 TSB, with or without 0.2% or 2% L-arabinose added, in 50 mL screw-cap conical centrifuge tubes (CellPro). The tubes were incubated for 24 hr at 25°C on a GyroMini nutating mixer (LabNet International, Inc.) at 24 rpm until the bacterial culture reached logarithmic growth. Cultures were then pelleted, the supernatant removed to a new 50 mL tube, and the pH was measured.
3
Salmonella Typhimurium Biofilm Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
S. Typhimurium biofilms were grown as follows: Overnight (O/N) cultures were grown in TSB at 37°C with aeration. These were normalized to OD600 = 0.8, diluted 1:10 into biofilm growth media (TSB diluted 1:20 or M9 +/- experimental conditions), and 0.1 mL was dispensed in triplicate into non-treated polystyrene 96-well plates (Corning). The plates were incubated for 24 hours (hr) at 25°C on a GyroMini nutating mixer (LabNet International, Inc.) at 24 rpm. If incubation longer than 24 hr was required, media was changed daily.
Attached biofilms of S. Typhimurium were then washed twice in ddH2O, heat fixed for 1 hr at 60°C, and stained with 0.33% crystal violet for 5 minutes (min). After two subsequent washes in ddH2O, the dye was released using 33% acetic acid, and the optical density was measured at 570 nm (OD570) in a SpectraMax Spectrophotmeter with SoftMax Pro software (Molecular Devices) to determine the amount of dye retained, which correlates to the amount of biofilm present. All biofilm experiments were performed in triplicate.
+ Expand
S. Typhimurium biofilms were grown as follows: Overnight (O/N) cultures were grown in TSB at 37°C with aeration. These were normalized to OD600 = 0.8, diluted 1:10 into biofilm growth media (TSB diluted 1:20 or M9 +/- experimental conditions), and 0.1 mL was dispensed in triplicate into non-treated polystyrene 96-well plates (Corning). The plates were incubated for 24 hours (hr) at 25°C on a GyroMini nutating mixer (LabNet International, Inc.) at 24 rpm. If incubation longer than 24 hr was required, media was changed daily.
Attached biofilms of S. Typhimurium were then washed twice in ddH2O, heat fixed for 1 hr at 60°C, and stained with 0.33% crystal violet for 5 minutes (min). After two subsequent washes in ddH2O, the dye was released using 33% acetic acid, and the optical density was measured at 570 nm (OD570) in a SpectraMax Spectrophotmeter with SoftMax Pro software (Molecular Devices) to determine the amount of dye retained, which correlates to the amount of biofilm present. All biofilm experiments were performed in triplicate.
4
Cellular Uptake Kinetics of Extracellular Vesicles
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were resuspended in 24 animals’ worth of EVs: either 0.5 ml PKH67-labeled EVs or 0.5 ml unlabeled EVs in CMF-B, or 0.5 ml control (no EV) solution. After resuspension, cells were incubated for 0, 1, 2, or 6 hr protected from the light and while rotating at 24 rpm on a GyroMini nutating mixer (Labnet, S0500) at RT. 45 min before the incubation with EVs was done, 50 μg/ml of Hoechst 33342 were added to label nuclei. Cells were centrifuged at 200 × g for 5 min and resuspended in 0.5 ml of CMF-B with Hoechst and propidium iodide (PI, 1 μg/ml). 0 hr samples were processed with 2 hr samples, but cells were centrifuged immediately after resuspension to remove EVs, then incubated 1X CMF-B only until Hoechst addition. Control samples for flow cytometry (unstained, unstained cells 2 hr after PKH67-labeled EV uptake, Hoechst, PI only, double stained, and x-irradiated) were prepared in parallel with the experimental samples.
5
Growth Curve Assay for Salmonella Typhimurium
Check if the same lab product or an alternative is used in the 5 most similar protocols
S. Typhimurium growth curves were performed as follows: O/N cultures were grown in TSB at 37°C with aeration. These were normalized to OD600 = 0.8, diluted 1:10 into 4 mL 1:20 TSB, with or without 0.2% or 2% L-arabinose added, in 5 mL screw-cap tubes (VWR). The tubes were incubated for 24 hr at 25°C on a GyroMini nutating mixer (LabNet International, Inc.) at 24 rpm. Aliquots were removed at designated time points, serial diluted 1:10 and 10 µL drip plated onto LB agar and incubated O/N at 37°C for quantification of CFU/mL. Growth curves were done in triplicate as biological replicates and averaged together. Growth curves performed in M9 were also normalized to OD600 = 0.8, diluted 1:10 into 1 mL M9 with 5 mM or 40 mM L-arabinose, and 0.1 mL was dispensed in triplicate into non-treated polystyrene 96-well plates (Corning). The plates were incubated for 24 hours (hr) static at 25°C, and the optical density was measured every 30 minutes, shaking for 5 seconds before each read, at 600 nm (OD600) in a SpectraMax Spectrophotmeter with SoftMax Pro software (Molecular Devices).
+ Expand
S. Typhimurium growth curves were performed as follows: O/N cultures were grown in TSB at 37°C with aeration. These were normalized to OD600 = 0.8, diluted 1:10 into 4 mL 1:20 TSB, with or without 0.2% or 2% L-arabinose added, in 5 mL screw-cap tubes (VWR). The tubes were incubated for 24 hr at 25°C on a GyroMini nutating mixer (LabNet International, Inc.) at 24 rpm. Aliquots were removed at designated time points, serial diluted 1:10 and 10 µL drip plated onto LB agar and incubated O/N at 37°C for quantification of CFU/mL. Growth curves were done in triplicate as biological replicates and averaged together. Growth curves performed in M9 were also normalized to OD600 = 0.8, diluted 1:10 into 1 mL M9 with 5 mM or 40 mM L-arabinose, and 0.1 mL was dispensed in triplicate into non-treated polystyrene 96-well plates (Corning). The plates were incubated for 24 hours (hr) static at 25°C, and the optical density was measured every 30 minutes, shaking for 5 seconds before each read, at 600 nm (OD600) in a SpectraMax Spectrophotmeter with SoftMax Pro software (Molecular Devices).
6
Consistent Sleep Deprivation in Flies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Consistent sleep deprivation was achieved using gentle mechanical stimulation on a GyroMini Nutating Mixer (Labnet International, Inc. Edison, NJ, USA). Vials containing 30–50 flies were placed at an angled position from the vertical center in a larger beaker with a raised block at a fixed position protruding inside the beaker on the mini gyrator. Mixer rotation caused the vials to rotate within the beaker and then gently jump over the raised block, providing the flies with a startle movement every 2.5 s. The constant motion of the vials combined with the startle ensured consistent sleep deprivation, with no apparent injuries or increased mortality observed after 24 h of sleep deprivation. Sleep deprivation was performed in an incubator under 25 °C, 60–70% relative humidity, and 12:12 LD conditions. Non-sleep-deprived controls were housed in the same incubator.
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!