Improved Cell Counting Accuracy at Extremely Low Cell Concentrations with a New Autofocus Algorithm of the LUNA-FX7
Modern cell counters utilize automated methods for cell counting within specific concentration range. The upper limit is influenced by the ability to accurately distinguish cell boundaries at high concentrations, while factors like the camera’s field of view (FOV) determine the lower limit. Nonetheless, in cases where an image contains only a few identifiable cells or features, autofocus failure can impede cell counting performance resulting in a higher coefficient of variation (CV). This is typically a minor issue in routine cell culture processes with sufficient cell concentration. However, it becomes critical in situations where the cell sample contains extremely low concentration, such as in the initial seed culture stage of cell therapy product manufacturing. In such cases, obtaining sharp images at extremely low cell concentrations is essential for achieving accurate cell counting results. To address this issue, we have developed a new autofocusing algorithm that employs advanced techniques to accurately identify features in images. This technical note discusses the latest updates in the LUNA-FX7™ software version (ver 1.9.1 or higher), which has been optimized for precise autofocusing in extremely low concentrations. Moreover, it showcases the exceptional performance of the LUNA-FX7™ automated cell counter.
Autofocusing Process in the LUNA FX7™
The autofocusing algorithm employed by the LUNA-FX7™ system uses image contrast to adjust focus. The autofocusing process consists of a coarse scan followed by a fine scan, ensures optimal focusing performance in the LUNA-FX7™ system (Figure 1). It begins with a coarse scan, capturing images across a wide range of Z-planes to cover a broad depth of field. During this scan, the algorithm identifies an approximate focal plane. Subsequently, a fine scan is performed, focusing on narrower Z-planes to precisely determine the most accurate focal plane. This process guarantees optimal focusing performance in the LUNA-FX7™ system based on image features.
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