Tissue dissociation and cell harvesting are two principal applicationsfor enzymes in tissue culture research and cell biology studies. Despitethe widespread use of enzymes for these applications over the years, theirmechanisms of action in dissociation and harvesting are not well understood.As a result, the choice of one technique over another is often arbitraryand based more on past experience than on an understanding of why the methodworks and what modifications could lead to even better results.

The goal of a cell isolation procedure is to maximize the yield of functionallyviable, dissociated cells. There are nine primary parameters which affectthe outcome of any particular procedure:

1. Type of tissue
2. Species of origin
3. Age of the animal
4. Dissociation medium used
5. Enzyme(s) used
6. Impurities in any crude enzyme preparation used
7. Concentration(s) of enzyme(s) used
8. Temperature
9. Incubation times

The first three items generally are not a matter of choice. To achievesuitable results the other six variable conditions are best defined empirically.

Researchers searching the scientific literature for information on theideal enzymes and optimal conditions for tissue dissociation are often confrontedwith conflicting data. Much of the variation stems from the complex anddynamic nature of the extracellular matrix and from the historical use ofrelatively crude, undefined enzyme preparations for cell isolation applications.Also, the extracellular matrix is composed of a wide variety of proteins,glycoproteins, lipids and glycolipids, all of which can differ in abundancefrom species to species, tissue to tissue and with developmental age. Commonlyused crude enzyme preparations such as Pronase, NF 1:250 and collagenasecontain several proteases in variable concentrations, as well as a varietyof polysaccharidases, nucleases and lipases.

The Worthington Tissue Dissociation guide summarizes our knowledge ofhow these enzymes accomplish the "routine" operations of tissuedissociation and cell harvesting and describes standard lab procedures.A logical experimental approach for establishing a cell isolation protocolis offered.

Note: We have not limited the references listed to only those papersusing Worthington enzymes. Generally speaking, the tissue dissociation enzymesoffered by Worthington can be used interchangeably for most preparationscited.