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Microplate reader

Manufactured by Allsheng
Sourced in China

The Microplate reader is a laboratory instrument used to measure the absorbance or fluorescence of samples in multi-well microplates. It can be used to quantify a wide range of analytes, such as proteins, enzymes, and nucleic acids. The device is capable of reading multiple samples simultaneously, providing efficient and high-throughput analysis.

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30 protocols using microplate reader

1

Quercetin Cytotoxicity Assay in H2O2-Treated Cells

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The harvested cells were seeded into the 96-well plate at the density of 3 × 103 cells per well and maintained at 37°C with 5% CO2 overnight. When the confluence reached 80–90%, the cells were treated for 24 h with different concentration of quercetin (Que, 0, 12.5, 25, 50, and 100 μM) [20 (link)], followed by addition 200 μM H2O2 treatment. The cells were then treated with addition of 10 μl of cell counting kit-8 (CCK-8) solution (Solarbio) and cultured for 4 h. Then, the cells were subjected to a microplate reader (Allsheng) to detect the optical density of cell at 450 nm.
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2

Quantitative Adipogenesis Assay

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The differentiated 3T3-L1 cells were washed with PBS and fixed with 10% formaldehyde for 1 h at room temperature followed by washing with PBS and 60% isopropanol. The cells were subsequently stained with Oil Red O solution (Sigma-Aldrich) for 30 min at room temperature. After washing with 60% isopropanol and distilled water, red oil droplets were visualized using an Eclipse Ts2-FL microscope (Nikon) at 100× magnification. Lipid accumulation was also quantified at an optical density of 492 nm using a microplate reader (Allsheng) after the cells were eluted using 100% isopropanol.
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3

MTT Assay for Cell Viability

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The 3T3-L1 cells (1 × 10 6 cells/mL) were plated on a 96-well culture plate for 48 h and subjected to a 24-h treatment with KI 32-W or KI 35-W at 37°C. After the treatment, the cells were incubated with 0.5 mg/mL 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) solution for 4 h at 37°C. Afterward, the supernatants were removed and formazan dark purple products were dissolved by adding 100 μL dimethyl sulfoxide into each well. The absorbance was measured at an optical density of 570 nm using a microplate reader (Allsheng).
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4

Triglyceride Quantification in 3T3-L1 Cells

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Triglyceride content in 3T3-L1 cells after treatment with KI 32-W or KI 35-W was measured using a commercial triglyceride assay kit (Abcam). Briefly, after the cells were extensively washed with PBS, the cell pellets were resuspended in 5% NP-40 diluted with distilled water and heated at 100°C for 5 min for extracting lipids. The cell suspensions were then centrifuged at 13,000 × g for 2 min, and the supernatants were mixed with lipase for converting triglyceride into glycerol fol-lowed by the addition of a triglyceride probe and a triglyceride enzyme mix. After reaction at room temperature for 1 h, the absorbance was measured at 570 nm using a microplate reader (Allsheng) and triglyceride concentration was calculated based on a standard curve of triglyceride standards.
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5

Comprehensive Biomolecular Analysis Protocol

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Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D, E4) (Biolin Scientific AB/Q-Sense, Gothenburg, Sweden) was used for affinity analysis; Real-time quantitative PCR system (TL980) (Xi’an Tianlong Science and Technology Ltd., Xi,an, China) was applied for monitoring the richness of library; PCR (T960) (Heal Force Bio-meditech Holdings Limited, Shanghai, China) was used for amplifying; F-7000 fluorescence spectrophotometer (Hitachi, Tokyo, Japan) was used for fluorescence detection; Microplate reader (Flash spectrum, Shanghai, China) was used for recording the absorbance; Micronucleic acid quantifier 3000 (Hangzhou Allsheng Instruments Co, Ltd., Hangzhou, China) was applied for ssDNA quantitation.
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6

Cytotoxicity Assay of Neutrophils

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The cytotoxicity assay was conducted using the CyQUANT™ MTT Cell Proliferation Assay Kit from Invitrogen (Cat. No. V13154). Neutrophils were seeded at a concentration of 2 × 105 cells per milliliter in conical tubes. The control and antioxidant-supplemented neutrophils (GSH + NAC and ALL) were pre-incubated for 60 min with their respective media (see materials and methods 4.2). After the pre-incubation period, the neutrophils were washed twice with PBS 1× to remove the media. Then, 100 µL of freshly prepared RPMI 1640 media alone was added to each well of a Nunc Flat-bottom microplate from Thermo Fisher (Cat. No. 168055), with a concentration of 2 × 105 cells per well. Subsequently, 10 µL of the CyQUANT™ MTT reagent was added to each well. The negative control of RPMI 1640 + MTT was included, as indicated by the manufacturer’s protocol. The plate was incubated for 3 h at 37 °C. After the incubation period, the plate was centrifuged for 10 min at 4 °C and 2000 rpm. Then, 85 µL of the total volume was carefully removed, and 50 µL of dimethyl sulfoxide (DMSO) was added to each well. The plate was incubated for 15 min, and each well was resuspended prior to reading. Absorbance readings were performed using a microplate reader (ALLSHENG) at 540 nm.
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7

Antioxidant Capacity Evaluation in Cells

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To evaluate the antioxidant effect in both medium and neutrophils, we utilized the Total Antioxidant Capacity Assay Kit from Abcam (Cat. No. ab65329, Cambridge, UK) following the manufacturer’s protocol without the protein mask. The TAC was assessed in the culture media components: RPMI 1640 alone, AS alone, and RPMI 1640 + AS, following the manufacturer’s instructions for liquid samples. For the evaluation of TAC in neutrophils, we seeded 1 × 106 neutrophils and followed the same protocol described in the section of NETosis assay for the controls and the combination of antioxidants GSH + NAC and ALL. After inducing NETosis, in all cases, the cell suspensions were washed twice with cold PBS 1× to remove the media. Subsequently, the samples were resuspended in 100 µL of ddH2O containing 0.05% Triton X-100 (Cat. No. 85111, Thermo Fisher). The resulting homogenate was then incubated on ice for 10 min. After incubation, the samples were centrifuged at top speed for 5 min at 4 °C. The supernatant was collected, and the assessment was carried out according to the instructions provided in the kit. The output of the TAC assay was measured using a microplate reader (ALLSHENG, Hangzhou, China) at 570 nm.
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8

Nitrite and Nitrate Quantification in Neutrophils

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To evaluate nitrite + nitrate production in neutrophils, we used the Nitrate/nitrite Colorimetric Assay Kit from Cayman (Cat. No. 780001, Ann Arbor, USA). Neutrophils were seeded at a concentration of 1 × 106 cells per milliliter in 1.5 mL conical tubes. The samples were pre-incubated for 60 min with their respective media as described in the media preparation section for controls and the antioxidant combinations (GSH + NAC and ALL). After pre-incubation, the cell suspensions were washed twice with cold PBS 1×, and the protocol described for NETosis assay was conducted. Subsequently, the cell suspensions were sonicated with 5 cycles of 5 s with intervals of 5 s off. The cells were centrifugated at 14,000 rpm for 30 min, and the supernatant was collected. The nitrite/nitrate assay was conducted using a volume of 80 µL of the processed samples, following the manufacturer’s protocol provided with the kit. The absorbance was measured at 550 nm using a microplate reader (ALLSHENG).
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9

Antibacterial Activity of Electrospun Webs

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The qualitative antibacterial activity of electrospun webs were evaluated according to standard AATCC 147-1998 through a disc diffusion assay against E. coli and S. epidermidis. First, 1 mL of overnight-precultured bacterial inoculums in nutrient broth was transferred to a new 10 mL fresh broth and incubated for 5 h at 200 rpm at 37 °C. The cultures were then diluted to an OD600 of 0.7 corresponding to 3.5 × 109 and 1.8 × 109 CFU mL−1 for E. coli and S. epidermidis, respectively, via a microplate reader (Allsheng, Hangzhou, China). The diluted culture was then spread over the entire surface of the nutrient agar plate using a spreader. Electrospun webs were cut into 20 mm diameter discs and then placed on the inoculated nutritional agar plate’s surface. The antimicrobial activity of the electrospun webs were recorded in terms of inhibition zone diameter (mm) using a manual caliper after plates were incubated for 24 h at 37 °C. Experiments were replicated for each sample. For the morphology of bacteria, samples were observed by SEM. Bacteria cells were fixed by osmium tetroxide vapor (2% w/v) for 24 h.
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10

MTT Assay for Cell Viability

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1 × 104 cells/well CAL27 and SCC25 cells were inoculated onto the 96-well plates for 12 h. Next, 10 μL of MTT (0.5 mg/mL, Beyotime, Shanghai, China) was supplemented into per well. 4 h later, medium was replaced by 100 μL of dimethyl sulfoxide (Thermo Fisher Scientific, Waltham, MA, USA). The absorbance was assessed using a microplate reader (Allsheng, Hangzhou, China) at 570 nm.
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