An automated method for gridding and clustering-based segmentation of cDNA microarray images

The Stanford Microarray Database (SMD) (http://smd.stanford.edu) is a research tool for hundreds of Stanford researchers and their collaborators. In addition, SMD functions as a resource for the entire biological research community by providing unrestricted access to microarray data published by SMD users and by disseminating its source code. In addition to storing GenePix (Axon Instruments) and ScanAlyze output from spotted microarrays, SMD has recently added the ability to store, retrieve, display and analyze the complete raw data produced by several additional microarray platforms and image analysis software packages, so that we can also now accept data from Affymetrix GeneChips (MAS5/GCOS or dChip), Agilent Catalog or Custom arrays (using Agilent's Feature Extraction software) or data created by SpotReader (Niles Scientific). We have implemented software that allows us to accept MAGE-ML documents from array manufacturers and to submit MIAME-compliant data in MAGE-ML format directly to ArrayExpress and GEO, greatly increasing the ease with which data from SMD can be published adhering to accepted standards and also increasing the accessibility of published microarray data to the general public. We have introduced a new tool to facilitate data sharing among our users, so that datasets can be shared during, before or after the completion of data analysis. The latest version of the source code for the complete database package was released in November 2004 (http://smd.stanford.edu/download/), allowing researchers around the world to deploy their own installations of SMD.

BackgroundMicroarray image analysis processes scanned digital images of hybridized arrays to produce the input spot-level data for downstream analysis, so it can have a potentially large impact on those and subsequent analysis. Signal saturation is an optical effect that occurs when some pixel values for highly expressed genes or peptides exceed the upper detection threshold of the scanner software (216 - 1 = 65, 535 for 16-bit images). In practice, spots with a sizable number of saturated pixels are often flagged and discarded. Alternatively, the saturated values are used without adjustments for estimating spot intensities. The resulting expression data tend to be biased downwards and can distort high-level analysis that relies on these data. Hence, it is crucial to effectively correct for signal saturation.ResultsWe developed a flexible mixture model-based segmentation and spot intensity estimation procedure that accounts for saturated pixels by incorporating a censored component in the mixture model. As demonstrated with biological data and simulation, our method extends the dynamic range of expression data beyond the saturation threshold and is effective in correcting saturation-induced bias when the lost information is not tremendous. We further illustrate the impact of image processing on downstream classification, showing that the proposed method can increase diagnostic accuracy using data from a lymphoma cancer diagnosis study.ConclusionsThe presented method adjusts for signal saturation at the segmentation stage that identifies a pixel as part of the foreground, background or other. The cluster membership of a pixel can be altered versus treating saturated values as truly observed. Thus, the resulting spot intensity estimates may be more accurate than those obtained from existing methods that correct for saturation based on already segmented data. As a model-based segmentation method, our procedure is able to identify inner holes, fuzzy edges and blank spots that are common in microarray images. The approach is independent of microarray platform and applicable to both single- and dual-channel microarrays.

<p>Microarray technology allows the simultaneous monitoring of thousands of genes. Based on the gene expression measurements, microarray technology have proven powerful in gene expression profiling for discovering new types of diseases and for predicting the type of a disease. Gridding, segmentation and intensity extraction are the three important steps in microarray image analysis. Clustering algorithms have been used for microarray image segmentation with an advantage that they are not restricted to a particular shape and size for the spots. Instead of using single feature clustering algorithm, this paper presents multiple feature clustering algorithm with three features for each pixel such as pixel intensity, distance from the center of the spot and median of surrounding pixels. In all the traditional clustering algorithms, number of clusters and initial centroids are randomly selected and often specified by the user. In this paper, a new algorithm based on empirical mode decomposition algorithm for the histogram of the input image will generate the number of clusters and initial centroids required for clustering. It overcomes the shortage of random initialization in traditional clustering and achieves high computational speed by reducing the number of iterations. The experimental results show that multiple feature Fuzzy C-means has segmented the microarray image more accurately than other algorithms.</p>

A new method for DNA microarray image segmentation based on pattern recognition techniques is introduced. The method performs an unsupervised classification of pixels using a clustering algorithm, and a subsequent supervised classification of the resulting regions. Additional fine tuning includes detecting region edges and merging, and morphological operators to eliminate noise from the spots. The results obtained on various microarray images show that the proposed technique is quite promising for segmentation of DNA microarray images, obtaining a very high accuracy on background and noise separation.KeywordsDNA microarray imagessegmentationclusteringclassification

Regular gridding and segmentation for microarray images

Microarray technology has become a power tool in the field of bioinformatics. It is used to measure gene expression levels and similar to any other image capturing processes is prone to noise. There are different kinds of noise, during preparation, hybridization and scanning in microarray images which usually are modeled by Gaussian noise. Since introduction of wavelets in 1970s, many more forms and extensions of this transform have been developed and used, such as stationary wavelet transform (SWT), complex wavelet transform (CWT), curvelet transform (CURV) and contourlet transform (CNT). By developing of more sparse transforms, it is important to have a perspective of how efficient the transforms are in different applications, such as microarray image analysis. In this paper, we compare the efficiency of common sparse transforms including ordinary discrete wavelet transform (DWT), SWT, CWT, CURV, CNT, Contourlet-SD decomposition, steerable pyramid (STP) and shearlet transform (SHR) for microarray image denoising. Therefore after converting microarray image into x-let transform, BayesShrink method, soft and hard thresholding are used to perform denoising of these images. Both local and general thresholds are calculated for each subband in order to evaluate the effect of incorporating intrascale dependency on top of sparsity property in statistical modeling of x-let's coefficients. Our simulation results show that CWT and SHR outperforms the others when using global thresholding and SWT is the preferred transform when using local thresholding. Although STP and SHR have better performance for some criteria like structural similarity (SSIM) index, but CWT is faster.

Microarray images, which allow the analysis of hybridization expressions of genes, have been one of the most widely used high-throughput technologies with many different applications. Accurate and automatic microarray image analysis is very important since researchers trust the information provided in these experiments and construct conclusions based on the results produced by the software responsible in analyzing the hybridized arrays. Every microarray image contains thousands of spots, so how to do the spots finding in microarray images accurately and automatically is very meaningful. There are always some problems, such as rotation and distortion, in a microarray image caused by mechanical errors and/or optical errors in the system. All these problems will hinder "doi"ng analysis automatically. Early research has worked out several algorithms to deal with the rotation problem, but those algorithms can not handle microarray images with distortions. In this paper, we propose a robust spots finding method to deal with both rotation and/or distortion in microarray images. The proposed method provides automatic gridding and can handle a microarray image with different type of rotation (global or sub-array rotation) and optical distortions.

Aiming at the problem that traditional fuzzy C-means (FCM) clustering algorithm is susceptible to noise in processing noisy images, a noisy image segmentation method based on FCM wavelet domain feature enhancement is proposed. Firstly, the noise image is decomposed by two-dimensional wavelet. Secondly, the edge enhancement of the approximate coefficient is carried out, and the artificial fish swarm (AFS) optimization algorithm is used to process the threshold value of the detail coefficient, and the processed coefficient is reconstructed by wavelet transform. Finally, the reconstructed image is segmented by FCM algorithm. Five typical gray-scale images are selected by adding Gaussian noise and Salt& pepper noise, respectively, and segmented by various methods. The peak signal-to-noise ratio (PSNR) and error rate (MR) of segmented images are used as performance indexes. Experimental results show that compared with traditional FCM clustering algorithm segmentation method, particle swarm optimization (PSO) segmentation method and other methods, the indexes of image segmentation by the proposed method is greatly improved. It can be seen that the proposed segmentation method retains the texture information of image edge well, and its anti-noise performance and segmentation performance are improved.

The Fuzzy C Means (FCM) and Expectation Maximization (EM) algorithms are the most prevalent methods for automatic segmentation of MR brain images into three classes Gray Matter (GM), White Matter (WM) and Cerebrospinal Fluid (CSF). The major difficulties associated with these conventional methods for MR brain image segmentation are the Intensity Non-uniformity (INU) and noise. In this paper, EM and FCM with spatial information and bias correction are proposed to overcome these effects. The spatial information is incorporated by convolving the posterior probability during E-Step of the EM algorithm with mean filter. Also, a method of pixel re-labeling is included to improve the segmentation accuracy. The proposed method is validated by extensive experiments on both simulated and real brain images from standard database. Quantitative and qualitative results depict that the method is superior to the conventional methods by around 25% and over the state-of-the art method by 8%.

Bead encoding is a key problem central to all multiplexed bead-based microarrays. Most existing bead-based microarrays require complicated and costly bead fabrication and/or sophisticated bead decoding hardware to achieve high multiplexing. Recently, the development of spatial bead encoding techniques have opened up the possibility of using pattern matching and image processing to develop highly multiplexed, high-throughput bead-based arrays. In this thesis, we improve the existing spatial bead encoding scheme, and develop a pipeline of computational methods, which allows for automated spatial bead encoding. Six novel computational methods, that automate the proposed bead encoding scheme, are developed in this thesis. The proposed scheme improves the previously reported spatial encoding schemes by making them better suited for use in real world scenarios. In the spatial bead encoding scheme, a sequential bead deposition method is used to capture the identities of the beads in bright or dark field images of the array. While, a second fluorescence image of the array is used to quantify the target analyte concentrations associated with the beads. By aligning the two images using the patterns formed by the beads, both the bead identities (i.e. the target analyte associate with the bead) and the analyte concentrations are decoded. The six computational methods developed in this thesis, are grouped into three categories: bright and dark field image processing, fluorescence image processing, and bead pattern matching. These methods are developed such that they require minimal parameter tuning, and are able to deal with noisy images acquired in uncontrolled environments As part of the bright and dark field image processing we have developed two novel methods: a fully automatic method for detecting the underlying micro-well grid structure in the images, and an unsupervised learning based method to classify the micro-wells as either empty or containing a bead. We show the ability of the proposed methods to deal with extreme amounts of noise and distortions using multiple datasets. While, microarray image gridding is a well-studied problem in the context of fluorescence images of planar microarrays, to the best of our knowledge no methods have yet been developed for processing either bright or dark field images of bead-based microarrays. Unlike in bright field images, where all the beads are visible, in fluorescence images only those beads that are expressed in the sample are visible. Therefore, in fluorescence image analysis the first step is to detect the beads in the image. To this end, we have developed an unsupervised, simultaneous bead detection and segmentation method which uses the coherence in shape and size of the beads to circumvent the need for parameter optimization. The main challenge in this segmentation is the large dynamic range across which the beads are expressed. We have also developed a probabilistic non-linear gridding method that attempts to establish the micro-well grid structure from the detected beads. While similar gridding techniques have been developed in the past, we make two contributions in this regard: first, we put the gridding method in a probabilistic framework, and second, we show that in certain cases it is impossible to ascertain if the correct grid has been detected without using additional information about the physical properties of the array. The final step in the proposed bead encoding scheme is to establishing one-to-one correspondences between the beads detected in the bright field and fluorescence images. We have developed two methods for this matching. When the micro-well grid structure in the fluorescence image is successfully established, the grid information can be used to reduce the matching problem to a binary grid alignment problem. A grid matching method that is very fast and can handle a large number of beads (>50,000) has been developed for this purpose. Finally, to perform matching when the fluorescence image grid is not available, a novel point pattern matching method that is affine invariant and robust to small non-linear distortions has been developed. The point pattern matching method addresses the problem of matching large point sets in the presence of large amounts of outliers and occlusion. The proposed method is evaluated using several real and simulated datasets.

Image processing is an important stage of every microarray experiment. Reliability of this stage strongly influences the results of data analysis performed on extracted gene expressions. Multiple methods related to array recognition, spot segmentation and measurement extraction have emerged in this area over past several years. Currently there are various commercial and freeware packages available, which perform microarray image analysis. This paper attempts to review microarray image analysis as a whole and to make some experimental comparison of several computational schemes for signal segmentation and measurement extraction. Also we provide a detailed discussion of automated image quality control for use with microarray images.

Fuzzy c-means (FCM) is a popular clustering method for image segmentation. However, FCM has difficulties in handling artifacts in brain magnetic resonance imaging (MRI), especially when it comes to bias field and noise. We propose a novel multiple-surface-approximation-based FCM with interval membership method for simultaneous bias correction and segmentation of Brain MRI. First, multiple surface representation of bias field is embedded into FCM to estimate and correct bias field. Then memberships of the improved FCM are extended to intervals. After the extension, clustering centers of different MR brain tissues could be solved more properly by the proposed method. Moreover, the proposed method is less sensitive to noise by introducing effects of neighboring pixels. Experiments conducted on artificial images and synthetic and real clinical Brain MRI show that the proposed method is effective and obtains better results of both bias field correction and segmentation than comparing methods.

Image segmentation is an important research topic in image processing and computer vision community. In this paper, a new unsupervised method for MR brain image segmentation is proposed based on fuzzy c-means (FCM) and fuzzy connectedness. FCM is a widely used unsupervised clustering algorithm for pattern recognition and image processing problems. However, FCM does not consider the spatial coherence of images and is sensitive to noise. On the other hand, fuzzy connectedness method has achieved good performance for medical image segmentation. However, in the computation of fuzzy connectedness, one needs to select seeds manually which is elaborative and time-consuming. Our new method used FCM as the first step to select salient seeded points and then applied fuzzy connectedness algorithm based on those seeds. Thus our method achieved unsupervised automatic segmentation for brain MR images. Experiments on simulated and real data sets proved it is effective and robust to noise.

Fuzzy C-means (FCM) clustering method has been widely used in image segmentation that plays an important role in a variety of applications in image processing and computer vision systems, but the performance of FCM heavily relies on the initial cluster centers which are difficult to determine. To solve the problem, the paper proposes a new hybrid method for image segmentation, which first randomly generates a population of initial clustering solutions and then uses an evolutionary algorithm to search for better clustering solutions; at each iteration, the FCM method is applied on each initial clustering solution to produce its segmentation result. Among a set of popular evolutionary algorithms, we find that the biogeography-based optimization (BBO) metaheuristic exhibits good performance on the considered problem. Besides the basic BBO, we have also proposed a set of improved BBO versions in their combination with FCM for image segmentation. Computational experiments on a set of test images show that the proposed method has significant advantage over the basic FCM algorithm and those hybrid algorithms combining FCM with other evolutionary algorithms such as artificial bee colony and particle swarm optimization.

In this paper, we propose a method of image segmentation, which is based on superpixel and optimized spatial feature. In this paper, the superpixels are taken into account, which can reduce computational burden, and the result of over-segmentation can also be benefit for segmentation results. The main idea of this paper is based on the conventional fuzzy c-means (FCM). The conventional FCM has a better performance. However, it is sensitive to noise. In order to overcome this shortage, we incorporate spatial information of superpixels into the conventional FCM. In order to obtain the better performance, influential degree of spatial information is applied to the conventional FCM to improve segmentation performance Experimental results show that the proposed method achieves excellent performance.