The protein samples collected during dialysis and ultrafiltration were analyzed for their concentration using a BioTek Epoch Multi-Volume Spectrophotometer (Winooski, VT, USA). A UV-transparent 96-well plate was utilized. Samples were pipetted in triplicate in 100-µL increments into the well plate. The absorbance was run at 280 nm for the BSA and lysozyme samples. The urea samples were prepared with a QuantiChrom Urea Assay Kit provided from VWR and run at 520 nm in a standard polystyrene 96-well plate, both supplied through VWR.
Epoch multi volume spectrophotometer
Manufactured by Agilent Technologies
Sourced in United States
The Epoch Multi-Volume Spectrophotometer is a lab equipment designed to measure the absorbance of light in various liquid samples. It can analyze multiple samples simultaneously within a single instrument.
Automatically generated - may contain errors
Lab products found in correlation
Take3, by Agilent Technologies (2 mentions)
Cell counting kit 8 (cck8), by Apexbio (2 mentions)
Horseradish peroxidase conjugated goat anti mouse igg antibody, by Merck Group (1 mentions)
Nunc maxisorp, by Thermo Fisher Scientific (1 mentions)
Coomassie plus protein assay kit, by Thermo Fisher Scientific (1 mentions)
Quantikine elisa kit, by R&D Systems (1 mentions)
Incucyte live cell analysis system, by Sartorius (1 mentions)
Wizard genomic dna purification kit, by Promega (1 mentions)
7 protocols using epoch multi volume spectrophotometer
1
Protein Concentration Analysis via Spectrophotometry
2
Quantification of Cholera Toxin by ELISA
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Quantification of LT in the bacterial fractions was performed by capture ELISA as previously described [15 (link)]. Briefly, the microtiter plates (Nunc Maxisorp-Thermo Fisher Scientific, Waltham, Massachusetts, USA) were coated with rabbit anti-cholera toxin serum (titer equal to 1 × 106) diluted to 1:1000 in PBS 1x. After overnight incubation, twofold serial dilutions of the samples were placed on the plates, and the captured LT was detected by serial exposure to mouse anti-LT serum (titer = 5 × 104) and horseradish peroxidase-conjugated goat anti-mouse IgG antibodies (Sigma-Aldrich, St. Louis, MO, USA) diluted to 1:2500 and 1:5000, respectively. The reactions were read at 492 nm in an Epoch™ Multi-Volume Spectrophotometer (BioTek Instruments, Vermont, USA). The Bradford assay was carried out to measure total protein in the bacterial samples according to the manufacturer’s instructions (Coomassie Plus Protein Assay Kit, Thermo Fisher Scientific). Standard curves (regression analysis with R2 > 0.98), generated with the absorbance values versus concentrations of purified LT or bovine serum albumin (Pierce-Thermo Fisher Scientific), were used to determine LT and total protein concentrations, respectively, in the samples. Purified LT was obtained as previously reported [15 (link)].
3
Quantification of Active TGF-β1 in Cell Supernatants
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The human bioactive TGF-β1 concentration in cell culture supernatants was measured using the Quantikine ELISA Kit (R&D Systems, Minneapolis, MN, USA). To activate latent TGF-β1, cell culture supernatants were acidified with 100 μL of 1 M HCl and incubated at room temperature for 10 min. The samples were then neutralized with 100 μL of 0.5 M HEPES/1.2 M NaOH. The absorbance was measured at 450 nm using an Epoch Multi-Volume Spectrophotometer (BioTek Instruments, Winooski, VT, USA). The limit of sensitivity was 15.4 pg/mL. All samples were analyzed in sextuplicate.
4
Stability Assessment of DOX Nanoformulations
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The absorption spectra of the DOX nanoformulation were measured by EPOCH multivolume spectrophotometer (BioTek Instruments, USA) in regular intervals to determine the stability and quality of the NPs following the protocol mentioned earlier9 (link). After the formulation of DOX loaded PLGA-PVA- NP and CS-DS coated DOX loaded PLGA-PVA- NP, a spectral scan analysis was performed in the range of 300–900 nm wavelength. We continued observing the spectral analysis for 30 days after every 24 h. The characteristic peaks obtained for PLGA, PVA, Chitsoan, DOX loaded PLGA-PVA- NP and CS-DS coated DOX loaded PLGA-PVA- NP were plotted and compared to conclude the uniqueness of the formulation.
5
Colorectal Cancer Cell Viability Assay
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Cell viability was determined by Cell Counting Kit-8 (K1080, Apexbio Technology LLC, Houston, TX, USA). CRC were seeded into 96-well plates or 24-well plates for 24 h. The cells were treated with indicated doses of DIM, 5-FU, and the combination of DIM and 5-FU for 48 h. Then, 10% (v/v) CCK-8 solution was prepared and added to every well, and cells were plated into a cell incubator for 3 h [42 (link)]. Optical density was measured at 450 nm using Epoch™ Multi-Volume Spectrophotometer and Take3™ (BioTek, Winooski, VT, USA). Cell viability was normalized as a percentage of the negative controls treated with DMSO.
6
Measuring CRC Cell Growth and Viability
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For growth curve, cell viability was monitored by the IncuCyte live cell analysis system (Essen BioScience, Ann Arbor, MI, USA). To assess the cell proliferation, CRC cells were equally seeded into fresh 12-well plates at a density of 1 × 105 (link) cells. Cells were treated with different indicated concentrations of C.B CM. After placing the cell in a cell incubator, the IncuCyte live cell analysis system analyze the photographs at 9 random fields to obtain growth curve.
For IC50 calculation, cell viability was determined by Cell Counting Kit-8 (K1080, Apexbio Technology LLC, Houston, TX, USA). CRC cells were seeded into 96-well plates for 24 h. The cells were treated with indicated doses of C.B CM, 5-FU, and the combination of C.B CM and 5-FU for 48–72 h. Following that, 10% (v/v) CCK-8 solution was prepared and added to every well, and cells were plated into a cell incubator for 24 h. Optical density was measured at 450 nm using Epoch™ Multi-Volume Spectrophotometer and Take3™ (BioTek, Winooski, VT, USA). Cell viability was normalized as a percentage of the negative controls treated with culture medium. IC50 values were calculated by using linear regression equations obtained from the concentration vs percentage of viable cells.
For IC50 calculation, cell viability was determined by Cell Counting Kit-8 (K1080, Apexbio Technology LLC, Houston, TX, USA). CRC cells were seeded into 96-well plates for 24 h. The cells were treated with indicated doses of C.B CM, 5-FU, and the combination of C.B CM and 5-FU for 48–72 h. Following that, 10% (v/v) CCK-8 solution was prepared and added to every well, and cells were plated into a cell incubator for 24 h. Optical density was measured at 450 nm using Epoch™ Multi-Volume Spectrophotometer and Take3™ (BioTek, Winooski, VT, USA). Cell viability was normalized as a percentage of the negative controls treated with culture medium. IC50 values were calculated by using linear regression equations obtained from the concentration vs percentage of viable cells.
7
DNA Extraction from Pumpkin Leaves
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A Wizard Genomic DNA purification Kit (Promega Corporation Biotechnology, Madison, WI, USA) was used for extracting DNA from the leaves of all inbred pumpkin lines. The extracted DNA was treated with RNase and stored in a refrigerator at -20 °C. The DNA quality was assessed using 0.8% agarose gel and an Epoch Multi-Volume Spectrophotometer (Biotek, Winooski, VT, USA). Before conducting the HFO-TAG analysis, the DNA was diluted to a 25 ng/µL concentration.
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