Cell counting kit 8 cck 8 kit

Short-chain chitosan (SCS, degree of deacetylation > 90%) was obtained from Jinhu Crust Product Co., Ltd. Branched polyethylenimine (PEI, 25 kDa), lrgacure 2959, sodium alginate (SA), and glycidyl methacrylate (GMA) were purchased from Energy Chemical Co. (Shanghai, China). The polydeoxyribonucleotide (PDRN) was obtained from ReaLi Tide Biological Technology Co., Ltd. (Weihai, China). A living/dead-cell double-staining kit, cell counting kit-8 (CCK-8 kit), a PicoGreen dsDNA assay kit, thiazolyl blue tetrazolium bromide (MTT), and triton X-100 were obtained from Solarbio Science & Technology Ltd. Fetal bovine serum (FBS), Dulbecco’s Modified Eagle’s Medium (DMEM), penicillin–streptomycin, and trypsin were obtained from Gibco Life Technologies (Carlsbad, CA, USA). Escherichia coli (E. coli, ATCC 25922) and Staphylococcus aureus (S. aureus, ATCC 6538) were acquired from the American Type Culture Collection (ATCC).

Cell proliferation was assessed using the Cell Counting Kit-8 (CCK-8 kit) (CA1210, Solarbio). SCC9 cells were seeded in 96-well plates at a density of 7 × 10³ cells per well. Subsequently, 110 µl of medium with 10 µl of CCK-8 reagent was added, and the cells were incubated for 2 hours at both 0 and 24 hours. The absorbance at 450 nm was then measured.
The cell migration (without Matrigel) and invasion (with Matrigel) assay was performed using a transwell chamber (3464, corning). Transfected cells, at a concentration of 10 × 10₄ cells per well, were seeded into the upper chamber with serum-free medium. In the lower chamber, complete medium supplemented with 10% FBS was used. Following a 48-hour incubation period, cells remaining on the membrane of the upper chamber were meticulously removed. The chambers were then fixed with paraformaldehyde and stained with crystal violet. Subsequently, the chambers were photographed and the cells were counted under a microscope.
Cells were seeded in 6-well plates; upon over 90% confluency, a 200-microliter pipette tip was used to create a vertical wound in the center of each well. Images were taken at 0 and 48 hours after the wound was created.

NKP608 was purchased from Medchem Express (New Jersey, USA). RPMI-1640 was supplied from Hyclone Laboratories (Logan, UT, USA). Fetal bovine serum (FBS) was from Gibco Cell Culture (Carlsbad, CA, USA). Cell counting kit-8 (CCK8) kit was obtained from Beijing Solarbio Science and Technology (Beijing, China). Metrigel was supplied with BD Biosciences (MA, USA). Annexin V-fluorescein isothiocyanate/propidium iodide (Annexin V-FITC/PI) Apoptosis Detection kit was from Beijing 4A Biotech (Beijing, China). Radioimmunoprecipitation assay (RIPA) lysis buffer and bicinchoninic acid (BCA) assay was from cwbiotech (Beijing, China).
Antibodies to Active-Caspase-3, Wnt-3a, β-catenin, E-Cadherin, (vascular endothelial growth factor) VEGF were obtained from Cell Signaling Technology, Inc. (Danvers, MA, USA). Other antibodies to Cyclin D1, Bcl-2, Bax, GAPDH and peroxidase conjugated secondary antibodies were provided from Proteintech Group, Inc. (Wuhan, China). The enhanced chemiluminescence (ECL) detection system was obtained from Proteintech Group, Inc. (Wuhan, China).

Furosine was purchased from PolyPeptide (Strasbourg, France); the purity was above 95%. Dulbecco’s Modified Eagle Medium (DMEM), fetal bovine serum (FBS), and non-essential amino acids (NEAA) were obtained from GIBCO (Waltham, MA, USA), 1% penicillin/streptomycin was purchased from Thermo Fisher (Waltham, MA, USA). Lactoferrin was purchased from Sigma (St. Louis, MO, USA), with the purity of above 95%.
Cell counting kit-8 (CCK-8 kit) was purchased from Solarbio (Beijing, China), Elisa detection kits of T, FSH and LH in mice serum were purchased from Jiancheng (Nanjing, China). Hematoxylin-Eosin (HE) staining kit was purchased from Solarbio (Beijing, China).
Trizol buffer, RIPA lysate buffer (including proteases and PMSF), and protein concentration detection kit were obtained from Beyotime (Nanjing, China). Primers of GAPDH, Cep55, NF-κB, PI3K, Akt, FOX01 and TNF-α were synthesized from Sangon (Shanghai, China), reagents related with quantitative real time polymerase chain reaction (q-PCR) were purchased from Sangon. Primary antibodies of β-actin, Cep55, NF-κB, PI3K, p-PI3K, Akt, p-Akt, FOX01, p-FOX01, and TNF-α, as well as the secondary antibodies, were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Reagents related with western blotting were purchased from Solarbio. Enhanced chemiluminescence (ECL) reagent was purchased from Tanon (Shanghai, China).

Cell viability was assessed using the Cell Counting Kit8 (CCK-8) kit ((#CK04-100 T; Solarbio) according to the manufacturer’s recommendations. A 10 µL volume of CCK-8 solution was added to each well, and the cultures were incubated at 37 °C for 1 h. After measuring the absorbance at 450 nm using a microplate reader, the results were presented as the percentage of viable cells relative to the control group. Each experiment included six readings for each experimental condition.

BA (purity ≥ 98%) was purchased from Nanjing Chunqiu Biological Engineering Co., Ltd. (Nanjing, China). Zein (protein content 92%) was provided by Sigma–Aldrich, Inc. (St. Louis, MO, USA). The human hepatocellular carcinoma HepG2 cell line was obtained from Prof. Wang [13 ,14 (link)]. Phthalaldehyde, coomassie brilliant blue G-250, and potassium bromide (spectral grade) were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). Standard protein (14.4–97.4 kDa), horseradish peroxidase, and β- mercaptoethanol were purchased from Thermor Science & Technologies Co., Ltd. (Xi’an, China). Additionally, 8-Amino-1-naphthalene sulfonic acid (ANS) was purchased from J&K Scientific Co., Ltd. (Beijing, China). Folin-phenol reagent, dialysis bag (molecular interception capacity of 8000 kDa), and cell counting kit-8 (CCK-8 kit) were purchased from Solarbio Biotechnology Co., Ltd. (Beijing, China). All chemicals, reagents, and solvents were analytically pure or of a higher grade.