Cell dissociation in vitro In animal and plant tissue culture, which methods can be used to disaggregate (separate) intact tissue into single, viable cells for primary culture initiation?

Introduction / Context:
Establishing primary cultures from tissues requires breaking cell–cell and cell–matrix connections to yield viable single cells. This question tests knowledge of standard laboratory strategies for tissue disaggregation in cell and tissue culture.

Given Data / Assumptions:

• Tissue pieces contain cells joined by adhesion molecules and extracellular matrix.
• Goal is high viability and adequate yield of single cells.
• Common laboratory tools and reagents are available (pipettes, sieves, enzymes, chelators).

Concept / Approach:
Disaggregation typically combines mechanical force to reduce tissue size, enzymes to digest proteins or polysaccharides in the matrix, and chelating agents to lower divalent cations that stabilize adhesions. The optimal mix depends on tissue type (animal vs. plant) and desired cell phenotype.

Step-by-Step Solution:

Verification / Alternative check:
Protocols for hepatocytes, fibroblasts, neurons, and plant protoplasts consistently employ combinations of the three approaches for optimal yield and viability.

Why Other Options Are Wrong:

• Physical disruption only: often insufficient and harsher forces reduce viability.
• Enzymatic digestion only: slower and less effective without size reduction and chelation.
• Chelation only: weakens adhesions but rarely achieves full dissociation alone.
• Antibiotics: control contamination but do not dissociate tissue.

Common Pitfalls:
Over-digestion (loss of surface receptors), excessive shear stress, or skipping chelation leading to persistent clumping.

Final Answer:
All of the above