FiloDetect: automatic detection of filopodia from fluorescence microscopy images

Abstract

BackgroundFilopodia are small cellular projections that help cells to move through and sense their environment. Filopodia play crucial roles in processes such as development and wound-healing. Also, increases in filopodia number or size are characteristic of many invasive cancers and are correlated with increased rates of metastasis in mouse experiments. Thus, one possible route to developing anti-metastatic therapies is to target factors that influence the filopodia system. Filopodia can be detected by eye using confocal fluorescence microscopy, and they can be manually annotated in images to quantify filopodia parameters. Although this approach is accurate, it is slow, tedious and not entirely objective. Manual detection is a significant barrier to the discovery and quantification of new factors that influence the filopodia system.ResultsHere, we present FiloDetect, an automated tool for detecting, counting and measuring the length of filopodia in fluorescence microscopy images. The method first segments the cell from the background, using a modified triangle threshold method, and then extracts the filopodia using a series of morphological operations. We verified the accuracy of FiloDetect on Rat2 and B16F1 cell images from three different labs, showing that per-cell filopodia counts and length estimates are highly correlated with the manual annotations. We then used FiloDetect to assess the role of a lipid kinase on filopodia production in breast cancer cells. Experimental results show that PI4KIII β expression leads to an increase in filopodia number and length, suggesting that PI4KIII β is involved in driving filopodia production.ConclusionFiloDetect provides accurate and objective quantification of filopodia in microscopy images, and will enable large scale comparative studies to assess the effects of different genetic and chemical perturbations on filopodia production in different cell types, including cancer cell lines.

Dystroglycan depletion inhibits the functions of differentiated HL-60 cells

“…Immunofluorescence quantification was performed using ImageJ (NIH, Bethesda, USA). Filopodia size and number were detected using the software FiloDetect from images taken from fluorescence microscopy [13].…”

Automated analysis of filopodial length and spatially resolved protein concentration via adaptive shape tracking

“…, 2011; Nilufar et al , 2013; Hendricusdottir and Bergmann, 2014; Tsygankov et al , 2014; Tarnok et al , 2015). Similarly, image analysis tools are available to measure spatiotemporal protein concentration in cellular structures such as the leading edge (Machacek et al , 2009; Barry et al , 2015).…”

Distinct focal adhesion protein modules control different aspects of mechanotransduction

“…Flilpodia were quantified using the FiloDetect MATLAB script, developed and made freely available by Theodore Perkins' research group, University of Ottawa (Nilufar et al, 2013).…”