Automated difference image analysis of lamellar ruffling: effect of temperature change on human SH-SY5Y neuroblastoma cells

Quantitation of cell movement and lamellar extension is critical to the stu dy of growth factors, chemotactic agents and signaling cascades. Many studi es are conducted by examining the size or number of lamellae in static imag es of cells. However, these methods do not quantify lamellar behavior over time and may overlook important changes in lamellar function. Most presentl y available methods for analyzing dynamic aspects of lamellar function exam ine changes in a cell's 2-dimensional perimeter and are best suited to the analysis of flattened lamellae. However, some cells generate 3-dimensional lamellar ruffles whose behavior is not readily detected using these methods . In the present study we analyze temperature-dependent ruffling of human S H-SY5Y neuroblastoma cells using automated digital subtraction of time-laps e images and quantitation of the resultant 'difference' image, and compare results obtained using this and other approaches. We report that ruffling b ehavior of SH-SY5Y cells is measurably altered by temperature changes of as little as 1 degreesC, and that these changes are best detected using diffe rence image analysis. Our studies indicate that temperature is a critical v ariable in studies of SH-SY5Y behavior and that difference image analyses m ay be an important complement to other methods in the study of lamellar ruf fling. (C) 2000 Elsevier Science B.V. All rights reserved.