IMAGE-ANALYSIS ASSESSMENTS OF PERFLUOROCARBON-ENHANCED AND SURFACTANT-ENHANCED PROTOPLAST DIVISION

Image analysis has been used to assess the growth of cell suspension-d erived protoplasts of Petunia hybrida cv. Comanche at an interface bet ween aqueous culture medium (KM8P), supplemented with 0.01% (w/v) Plur onic F-68, and oxygenated (10 mbar; 10 min) perfluorodecalin. Protopla sts synthesised a new cell wall and entered normal mitotic division wh ich was sustainable to the cell colony/callus stage. This process was accentuated by the collective and additive effects of oxygen, perfluor odecalin and surfactant media supplements. The mean area (mm(3)) of pr otoplast-derived cell colonies after 68 days of growth was increased 3 5 fold over control (media alone) in the presence of these combined tr eatments. The new cultural regime, leading to improved cell throughput from protoplasts, is discussed primarily in relation to the role of p erfluorodecalin as a gas carrier and possible effects of Pluronic F-68 in stimulating cellular uptake of nutrients and/or growth regulators. Image analysis provides a novel and accurate approach to quantifying cell growth responses.