Spectra max 250 spectrophotometer

Biofilm formation was determined using the previously described method with modifications (24 (link)). Bacterial strains were inoculated into the MB medium and incubated for 24 h at 30°C. Bacterial cells were collected by centrifugation and resuspended in fresh MB medium to an OD₆₀₀ of 0.2. To form a mixed-biofilm, the cell suspensions of two different strains were mixed at a 1:1 OD₆₀₀ ratio. Each cell suspension was diluted to an OD₆₀₀ of 0.01 in fresh MB medium. Flat-bottom 96-well polystyrene microtiter plates (Thermo Fisher Scientific Inc. IL, USA) were used to examine biofilm formation. An aliquot of 100 μL of the diluted cell suspension was added to each well. After being incubated at 30°C for 48 h, 25 μL of a 0.1% crystal violet solution was added to each well. The plates were incubated at room temperature for 15 min and rinsed twice with sterile water. Crystal violet was dissolved in 100 μL of 99.5% ethanol, and biofilm formation was analyzed at 595 nm using a Spectra Max 250 spectrophotometer (Molecular Devices, CA, USA).

To assess the cytotoxicity
of the compounds being studied, the XTT Cell Viability Assay kit (Biological
Industries, USA) was utilized as per the manufacturer’s guidelines.
In brief, Vero cells were treated with varying concentrations of the
conjugates under study for 3 days at 37 °C. Postincubation, the
medium was discarded, and 100 μL of new medium was added to
the cells. Subsequently, 50 μL of the activated XTT solution
(2,3-bis(2-methoxy-4-nitro-5-sulphenyl)-(2H)-tetrazolium-5-carboxanilide)
was added, followed by a further incubation for 2 h at 37 °C.
Absorbance was measured at a wavelength of 450 nm using a Spectra
MAX 250 spectrophotometer (Molecular Devices, USA). The results were
expressed as a percentage, calculated as the ratio of the signal from
the tested sample to that from the control sample (cells treated with
solvent) multiplied by 100%.

Transfected MDA-MB-231 and MCF-7 cells were seeded at a density of 5 × 10³ cells per well into 96-well plates and incubated at 37 °C for 24 h. Cell viability was assessed at 24, 48, 72 and 96 h using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay [31 (link)]. Absorbance values were determined at 570 nm (SpectraMax 250 spectrophotometer; Molecular Devices, Sunnyvale, CA, USA).

Cell Counting Kit 8 (CCK-8; Beyotime Biotech) assays were used to measure cell viability from OD₄₅₀ absorbance readings (Spectra Max 250 spectrophotometer; Molecular Devices, California, USA). After exponential phase, cells were seeded into 96-well plates, 10 μl CCK-8 reagent was added to each well at 6, 24, 48, 72, and 96 h [27 (link)]. After 1.5 h of incubation, absorbance was measured using a microplate reader. Each experiment was performed independently in triplicate.
Colony formation assays were performed to determine the colony numbers. Briefly, HCC cells were seeded into 6-well plates and maintained for 14 days. Emergent cell colonies were then fixed and incubated with 0.1% crystal violet and 20% methanol solution for 10 min. Visible colonies were photographed and counted using a microscope (Nikon, Tokyo, Japan). All experiments were performed at least thrice.

The proliferation rate of cells was measured using the CCK-8 assay Kit (CCK-8, Beyotime Institute of Biotechnology, Shanghai, China). Cells were seeded into 96-well plates at the indicated density and cultured in complete medium. The absorbance was measured at 450 nM using a SpectraMax 250 spectrophotometer (Molecular Devices, USA). The EdU assay (RiboBio, Guangzhou, China) was used to determine the cell proliferation capacity. The 97H cells were seeded into 96-well plates at a density of 1×10⁴ cells per well and cultured overnight. All operations were performed based on the kit instructions. Fluorescence microscopy was used to observe and analyze the number of EdU-positive cells.