Arcegel Matrix Gel Application - Hepatocellular Carcinoma Organoid

Arcegel Matrix Gel Application - Hepatocellular Carcinoma Organoid


According to data from Our World in Data, as of 2019, the mortality rate of liver cancer ranks fourth among all cancer deaths in China. According to statistics from the International Agency for Research on Cancer (IARC) of the World Health Organization, there were 19.29 million new cancer cases worldwide in 2020, with 4.57 million new cases in China. Among them, liver cancer accounted for 910,000 new cases globally, with approximately 410,000 cases in China, accounting for 45% of global cases. In a study published by Broutier et al. from the University of Cambridge in 2017, a novel organoid closely resembling primary liver cancer (PLC) was developed. This achievement greatly enhances the understanding of the pathophysiology of liver cancer and enables significant advancements in drug screening. The research findings were published in the journal Nature Medicine.

 

Figure 1 Sourced from Our World in Data

Organoids are 3D cell cultures derived from stem cells, progenitor cells, tumor cells, etc., cultivated in vitro with certain spatial structural organization resembling tissues. As disease models, organoids offer a closer approximation to the in vivo environment in elucidating disease progression, homeostasis, and pathogenesis compared to traditional 2D disease models. Organoids are primarily constructed from adult stem cells or tumor samples under suitable culture conditions. Mixing with extracellular matrix (ECM), or matrix gel, is one of the key steps to provide a 3D growth scaffold for organoids. The main components of matrix gel include laminin, type IV collagen, heparan sulfate proteoglycans (HSPGs), and nidogen, along with growth factors, tissue plasminogen activator, and other growth factors naturally present in EHS tumor. It provides support, tensile strength, and scaffold support for tissues and cells. Arcegel matrix gel developed and produced by Yeksa Biotechnology is free from LDEV (lactate dehydrogenase-elevating virus), with an ultra-low endotoxin content, and has undergone thorough mycoplasma testing to ensure no mycoplasma contamination, including different types of matrix gel such as basic concentration, high concentration, low growth factor, etc.

Application Direction

Product Features

 High safety: Free from LDEV (Lactate Dehydrogenase-Elevating Virus)

Diverse concentration range: Concentration ranges from 8 to 20 mg/mL

Good batch stability: Strict production quality inspection process ensures stable performance between batches

Low endotoxin: Endotoxin content < 8 EU/mL

Contaminant detection: Tested free from mycoplasma, bacteria, and fungal residues

High yield: Production exceeding 50L/month

Good compatibility: Compatible with any type of cell culture medium

Hepatocellular carcinoma organoid construction

Experimental Procedure

1. Place the pipette tips on ice for pre-cooling, thaw the matrix gel on ice or at 4°C, and pre-warm the 24-well cell culture plate at 37°C.

2. Harvest HepG2 cells in logarithmic growth phase, gently wash with PBS, and digest cells with trypsin.

3. After digestion, prepare and collect single-cell suspension, centrifuge at 1500 rpm for 3 minutes, discard the supernatant, and resuspend cells in DMEM complete medium, adjusting the cell concentration to 1.0×106 cells/mL.

4. Cell seeding: Mix thawed matrix gel with the adjusted concentration of single-cell suspension at a ratio of 1:1 on ice, gently pipette to mix. Using pre-cooled 200μL pipette tips, pipette 40-50μL of the mixed single-cell suspension vertically into the pre-cooled 24-well plate to form arched cell droplets.

5. After stabilizing for 30 minutes in a cell culture incubator at 37°C, 5% CO2 saturation humidity, add 500-2mL of hepatocellular carcinoma organoid complete medium to each well for continued culture.

6. Observe and photograph daily, and uniform hepatocellular carcinoma organoids can be observed approximately 5-7 days later.

 

Experimental Results

Figure 2 Results of organoid culture from HepG2 cell line after 5 days of cultivation.

FAQ

1. What precautions should be taken when handling matrix gel?

All operations should be carried out in a sterile environment, and pre-cooled pipettes should be used to ensure that the matrix gel is in a uniform slurry state.

2. How to aliquot, freeze, and use matrix gel? 

After thawing, Arcegel matrix gel can be aliquoted into multiple small tubes, all aliquoting should be done using pre-cooled cryovials, quickly frozen, and stored to avoid repeated freeze-thaw cycles.

3. What are the volumes of matrix gel and culture medium required for culturing organoids in different well plates? Organoids are commonly cultured in 24-well plates with 50μl/droplet of matrix gel, followed by adding 500~800μl of culture medium to cover the droplets. In 96-well plates, 10μl/droplet of matrix gel is used, followed by adding 200μl of culture medium to cover the droplets. In 6-well plates, multiple 50μl droplets can be seeded per well, followed by adding 2~3ml of culture medium to cover the droplets.

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