Tissue dissociation challenge: During isolation of free cells and cell aggregates from solid organs, what is the major technical problem to overcome?

Difficulty: Easy

Correct Answer: Releasing the cells from their supporting extracellular matrix without damaging viability

Explanation:


Introduction / Context:
Preparing primary cell suspensions from tissues (liver, kidney, tumors) requires breaking cell–cell junctions and cell–matrix adhesions while preserving cell viability and functionality. This is foundational for primary culture, single-cell analyses, and regenerative studies.


Given Data / Assumptions:

  • Solid organs contain extensive extracellular matrix (ECM) and tight junctional complexes.
  • Target outcome is viable single cells or small aggregates.


Concept / Approach:
The major challenge is to release cells from the ECM and from each other without excessive shear or proteolytic damage. Enzymes such as collagenase, dispase, elastase, and trypsin, plus chelators (EDTA/EGTA) and mechanical trituration, are used carefully to disrupt ECM and adhesions. Over-digestion or harsh shear compromises viability and surface receptors.


Step-by-Step Solution:

Define the goal: obtain free cells/aggregates while preserving viability.Identify the barrier: strong ECM and adhesion molecules (collagen, laminin, integrins, cadherins).Apply controlled enzymatic and mechanical methods to release cells from the matrix.


Verification / Alternative check:
Quality checks include trypan blue viability, yield counts, and functional assays; optimizing enzyme concentration, temperature, and time improves outcomes.


Why Other Options Are Wrong:

  • “Inhibiting” detachment contradicts the goal.
  • “Disintegrating cells” destroys the preparation.
  • Avoiding enzymes/chelators entirely is unrealistic for most tissues.
  • Removing organelles is neither feasible nor relevant.


Common Pitfalls:
Overexposure to proteases, insufficient washing, and excessive shear leading to low viability or altered phenotype; ignoring tissue-specific protocols.


Final Answer:
Releasing the cells from their supporting extracellular matrix without damaging viability

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