Cck 8 solution

To detect CCK-8, 10³ HCC cells were inoculated into 96-well plates and treated with 10 μl CCK-8 solution (RiboBio, China) at 0, 24, 48, and 72 h, respectively. According to the instructions of the manufacturer (Synergy, USA), cell absorbance was measured at the corresponding time point of 450 nm using a microplate reading element.
With the help of a Cell-Light EdU DNA Cell Proliferation Kit (RiboBio, China), we conducted a 5-ethynyl-2′-deoxyuridine (EdU) assay to evaluate cell proliferation. We put 5 × 10⁴ HCC cells into 24-well plates, then cultured the cells for 24 h, fixed the cell lines with 4% paraformaldehyde after incubation with 50 L for 2 H/L EdU solution. After the agreement with the manufacturer, we put the Apollo dye solution of the cell line and Hearst seal separately. EdU cell lines were collected and calculated under an Olympus FSX100 microscope (Olympus, Japan).

For CCK8 experiments, CRC cells were cocultured with TAMs supernatant containing sh-NC or sh-APOC1. Ten microliters of CCK8 solution (RiboBio, China) were applied at 0 hours, 24 hours, 48 hours, and 72 hours after the cocultured tumor cells were implanted in 96 wells. 4 hours after adding the CCK8 solution, analyses were carried out using a microplate reading element at 450 nm in accordance with the manufacturer's instructions (Synergy4, USA).

Cells were seeded at 2 × 10³ cells/well into 96-well plates, and the cells were routinely cultured. Each well was added with 10 μL of CCK-8 solution (RiboBio, Guangzhou, China) at 24, 48, 72, and 96 h, respectively, followed by the incubation for 1 h. Subsequently, the absorbance at 450 nm was determined by a synergy microplate reader (BioTek, Winooski, VT, USA).

Cancer cells were treated with supernatant derived from TAM, which received PBS or αAPOE treatment. Then cancer cells were seeded in 96 wells and then administrated with 10 μl of CCK8 solution (Ribobio, China) when cultured at 0h, 24h, 48h, and 72h, separately. Subsequently, cells absorbance at the respective time received the analysis at 450 nM by microplate reading element by conforming to the producer's instructions (Synergy4, USA).

In the clone forming experiments, HCC cells under transfection received the seeding inside 6-well plates at 1000 cells per well density. After 10 days, the cells received the fixing based on the use of methanol, followed by the staining with GIMSA. Eventually, colonies received the imaging and counting. For CCK8 assay, HCC cells were seeded in 96-well plates at 4000 cells per well. Seeded cells received 10 μl of CCK-8 solution (RiboBio, China) at 0 h, 24 h, 48 h, 72 h, and 96 h. Subsequently, cell absorbance at 450 nm was analysed at the respective times with a microplate reader in accordance with the manufacturer’s instructions (Synergy4, USA). Using a Cell-Light EdU DNA Cell Proliferation Kit (RiboBio, China), an EdU experiment was performed to assess the proliferation of cells. HCC cells were plated in 24-well plates and cultured for 24 h. The mentioned cell lines were fixed with 4% paraformaldehyde after incubation with 50 mM EdU solution for 2 h. Next, in accordance with the manufacturer’s protocol, cell lines underwent a sealing process with Apollo Dye Solution and Hoechst separately. Under an Olympus FSX100 microscope (Olympus, Japan), the EdU cell lines were imaged and counted.

In the clone forming experiment, CRC cells were seeded in 6-well plates at a density of 1000 cells/well. After 10 days, the cells were fixed using methanol (4%; Wobixin Inc., China), followed by staining with Giemsa (Sigma-Aldrich, USA), and the colonies were then imaged and counted.
To perform the CCK-8 assay, we seeded 10³ CRC cells in 96-well plates and then treated them with 10 μL of CCK-8 solution (RiboBio, China) at 0 h, 24 h, 48 h, 72 h, and 96 h of culturing. The cell absorbance was measured at the respective time points at 450 nm using a microplate reading element, according to the manufacturer's instructions (Synergy, USA).
Using Cell-Light 5-ethynyl-2'-deoxyuridine (EdU) DNA Cell Proliferation Kit (RiboBio, China), we performed the EdU experiment to assess cell proliferation. We plated 5 × 10⁴ CRC cells in 24-well plates, and the cells were then cultured for 24 h. The cell lines were fixed with 4% paraformaldehyde after incubating them with 50 mmol/L EdU solution for 2 h. Following the manufacturer's protocol, we treated the cell lines with Apollo Dye Solution and Hoechst seal, respectively. EdU cell lines were captured and counted under Olympus FSX100 microscope (Olympus, Japan).