Solid KHT tumor dispersal for flow cytometric cell kinetic analysis. [Bacteria]
A bacterial neutral protease was used to disperse KHT solid tumors into single cell suspensions suitable for routine cell kinetic analysis by flow cytometry and for clonogenic cell survival. Neutral protease disaggregation under conditions which would be suitable for routine tumor dispersal was compared with a trypsin/DNase procedure. Cell yield, clonogenic cell survival, DNA distributions of untreated and drug-perturbed tumors, rates of radioactive precursor incorporation during the cell cycle, and preferential cell cycle phase-specific cell loss were investigated. Tumors dispersed with neutral protease yielded approximately four times more cells than those dispersed with trypsin/DNase and approximately a 1.5-fold higher plating efficiency in a semisolid agar system. Quantitative analysis of DNA distributions obtained from untreated and cytosine-arabinoside-perturbed tumors produced similar results with both dispersal procedures. The rates of incorporation of tritiated thymidine during the cell cycle were also similar with neutral protease and trypsin/DNase dispersal. Preferential phase-specific cell loss was not obseved with either technique. We find that neutral protease provides good single cell suspensions of the KHT tumor for cell survival measurements and for cell kinetic analysis of drug-induced perturbations by flow cytometry. In addition, the high cell yields facilitate electronic cell sorting where large numbers of cells are often required.
- Research Organization:
- Lawrence Livermore National Lab., CA
- DOE Contract Number:
- W-7405-ENG-48
- OSTI ID:
- 5177131
- Journal Information:
- Cytometry (Baltimore); (United States), Vol. 2:2
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ENZYMES
BIOCHEMICAL REACTION KINETICS
NEOPLASMS
DISPERSIONS
TUMOR CELLS
CELL FLOW SYSTEMS
BACTERIA
CELL CYCLE
DNA
THYMIDINE
TRITIUM COMPOUNDS
ANIMAL CELLS
AZINES
DISEASES
HETEROCYCLIC COMPOUNDS
KINETICS
LABELLED COMPOUNDS
MICROORGANISMS
NUCLEIC ACIDS
NUCLEOSIDES
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
PYRIMIDINES
REACTION KINETICS
RIBOSIDES
550301* - Cytology- Tracer Techniques