Accutase Cell Detachment Solution: Gentle on Cells, Powerful in Performance

During adherent cell culture, cell dissociation is an essential step that directly impacts cell health and experimental outcomes. The trypsin/EDTA method is a classic and widely used approach known for its high efficiency. However, it also presents certain limitations. When gentle dissociation is needed to preserve cell viability, function, or surface markers, or when working with sensitive or specialized cell types, the following challenges often arise:

1.Cell surface proteins and extracellular matrix components are susceptible to damage.

2.Excessive enzymatic digestion can result in the loss of surface markers.

3.Cell viability is reduced.

To address these challenges in cell dissociation, this issue of Cell Culture Academy introduces a gentle and highly efficient reagent, Accutase Cell Detachment Solution, offering a new option for handling difficult-to-dissociate and sensitive cell types.

Ⅰ. What is Accutase?

Accutase is a non-animal-derived cell dissociation enzyme solution that contains no components of mammalian or bacterial origin. It is formulated with a blend of proteolytic and collagenolytic enzymes that replicate the synergistic activity of natural digestive fluids.

Accutase gently hydrolyzes intercellular adhesion molecules (such as cadherins and integrin ligands) and extracellular matrix proteins (such as fibronectin and laminin). It also breaks protein bonds between cells and the culture surface, enabling efficient and gentle cell detachment.

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Ⅱ. Four Features of Accutase

Thanks to its unique enzymatic properties, Accutase offers the following four key features:

1. Gentle Digestion

Accutase is a mild cell dissociation enzyme mixture that cleaves cell-surface proteins minimally, reducing damage to the cell membrane and lowering cell mortality. It is especially suitable for highly sensitive cell types, such as human or mouse embryonic stem cells.

2. Efficient Separation

Although digestion with Accutase is gentle, it remains highly efficient. Most adherent cells can be detached within minutes. Typically, additional washing or neutralization is not required, saving experimental time and improving passage efficiency. For cells that are slow to digest using traditional methods, Accutase often delivers ideal results.

3. Preservation of Cell Viability

Accutase minimizes cell stress and damage during dissociation, maintaining high cell viability and promoting faster recovery to normal growth. In cryopreservation and thawing experiments, cells treated with Accutase also demonstrate better post-thaw recovery.

4. Protection of Surface Antigens

Based on its unique enzymatic profile, Accutase minimizes excessive degradation of cell-surface proteins and antigens, preserving their integrity. This ensures reliable results in downstream applications such as flow cytometry, cell sorting, and immunophenotyping.

Ⅲ. Applicable Cell Types

Accutase is highly versatile and can be used with most mammalian cell lines and primary cells. It is also effective for certain insect cells. The following are examples of situations where Accutase offers particular advantages:

1. Human Pluripotent Stem Cells (hPSCs)

Human pluripotent stem cells include embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). These cells are extremely sensitive to dissociation conditions, and trypsin may induce spontaneous differentiation under certain. Accutase’s gentle dissociation preserves cell-surface molecules, maintains stemness and pluripotency, and significantly improves cell survival. It is especially recommended when single-cell dissociation is required.

2. Adult Stem Cells

Adult stem cells, including adherent and suspension types such as neural stem cells (NSCs), dental pulp stem cells (DPSCs), and mesenchymal stem cells (MSCs), are sensitive to enzymatic digestion. Accutase minimizes dissociation-induced damage, preserves cellular function and differentiation potential, and is suitable for downstream applications. Digestion time and temperature should be optimized based on cell type and culture conditions to achieve the best results.

3. Common Cell Lines

Accutase is suitable for most adherent cell lines, such as HeLa and HEK-293T. While these cells can also be digested with trypsin, Accutase better preserves cell-surface antigens. The resulting cells are ideal for flow cytometry, cell sorting, and immunophenotyping applications.

4. Primary Cells

Some primary cells, such as cardiac microvascular endothelial cells, are difficult to dissociate into high-quality single-cell suspensions using traditional methods. Accutase improves dissociation efficiency and enhances cell viability, making it an ideal choice for isolating these sensitive cells.

IV. Application Examples

Pricella® has introduced the gentle and efficient Accutase Cell Detachment Solution (Cat. No.: PB180201), validated across multiple cell types.

1.Rat Bone Marrow Mesenchymal Stem Cells

Figure 1. Rat bone marrow mesenchymal stem cells

 (Left: Pre-Digestion; Right: Post-Digestion)

Figure 2. Flow cytometry analysis of rat bone marrow mesenchymal stem cells (CD90+/CD44+)

Rat bone marrow mesenchymal stem cells were digested using Accutase Cell Detachment Solution (Cat. No.: PB180201). After digestion, cells were seeded into a 6-well plate at a 1:3 subculturing ratio. By day three, the cultures reached 100% confluence, with more than 3×106 cells and viability exceeding 92%. Flow cytometry analysis confirmed that CD90 and CD44 were expressed on 95% of the cells.

2.Rat Neural Stem Cells

Figure 3. Rat neural stem cells

(Left: Pre-Digestion; Middle: Post-Digestion; Right: Immunofluorescence Nestin)

Rat neural stem cells were dissociated using Accutase Cell Detachment Solution (Cat. No.: PB180201). Post-dissociation cell viability, assessed by Trypan Blue exclusion, exceeded 99%. Immunofluorescence staining with Nestin antibody confirmed that over 90% of the cells were positive.

To achieve optimal dissociation across different tissue or cell types, digestion time should be adjusted based on observed cellular response. During use, it is recommended to monitor the digestion endpoint under a microscope to ensure precise and controlled dissociation.