Andy’s Algorithms: new automated digital image analysis pipelines for FIJI

Quantification of cellular antigens and their interactions via antibody-based detection methods are widely used in scientific research. Accurate high-throughput quantitation of these assays using general image analysis software can be time consuming and challenging, particularly when attempted by users with limited image processing and analysis knowledge. To overcome this, we have designed Andy’s Algorithms, a series of automated image analysis pipelines for FIJI, that permits rapid, accurate and reproducible batch-processing of 3,3′-diaminobenzidine (DAB) immunohistochemistry, proximity ligation assays (PLAs) and other common assays. Andy’s Algorithms incorporates a step-by-step tutorial and optimization pipeline to make batch image analysis simple for the untrained user and adaptable across laboratories. Andy’s algorithms provide a simpler, faster, standardized work flow compared to existing programs, while offering equivalent performance and additional features, in a free to use open-source application of FIJI. Andy’s Algorithms are available at GitHub, publicly accessed at https://github.com/andlaw1841/Andy-s-Algorithm.

[1]  Andrew K. C. Wong,et al.  A new method for gray-level picture thresholding using the entropy of the histogram , 1985, Comput. Vis. Graph. Image Process..

[2]  Johan Hartman,et al.  Estrogen receptor beta in breast cancer—Diagnostic and therapeutic implications , 2009, Steroids.

[3]  Nikhil R. Pal,et al.  On minimum cross-entropy thresholding , 1996, Pattern Recognit..

[4]  J A Ramos-Vara,et al.  When Tissue Antigens and Antibodies Get Along , 2014, Veterinary pathology.

[5]  Sebastian Herzog,et al.  MCL-1 inhibition provides a new way to suppress breast cancer metastasis and increase sensitivity to dasatinib , 2016, Breast Cancer Research.

[6]  G. Zack,et al.  Automatic measurement of sister chromatid exchange frequency. , 1977, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[7]  U. Landegren,et al.  Direct observation of individual endogenous protein complexes in situ by proximity ligation , 2006, Nature Methods.

[8]  Vilppu J Tuominen,et al.  ImmunoRatio: a publicly available web application for quantitative image analysis of estrogen receptor (ER), progesterone receptor (PR), and Ki-67 , 2010, Breast Cancer Research.

[9]  C. M. van der Loos.  Multiple immunoenzyme staining: methods and visualizations for the observation with spectral imaging. , 2008, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[10]  C. Begley,et al.  Drug development: Raise standards for preclinical cancer research , 2012, Nature.

[11]  Herbert Herzog,et al.  Y4 receptor knockout rescues fertility in ob/ob mice. , 2002, Genes & development.

[12]  Bülent Sankur,et al.  Survey over image thresholding techniques and quantitative performance evaluation , 2004, J. Electronic Imaging.

[13]  Phil M E Waite,et al.  Axonal injury in children after motor vehicle crashes: extent, distribution, and size of axonal swellings using beta-APP immunohistochemistry. , 2002, Journal of neurotrauma.

[14]  Marcel E Dinger,et al.  Cyclin E2 is the predominant E-cyclin associated with NPAT in breast cancer cells , 2015, Cell Division.

[15]  Abhijit G. Shanbhag,et al.  Utilization of Information Measure as a Means of Image Thresholding , 1994, CVGIP Graph. Model. Image Process..

[16]  A. Ruifrok,et al.  Quantification of histochemical staining by color deconvolution. , 2001, Analytical and quantitative cytology and histology.

[17]  Jay Neitz,et al.  The genetics of normal and defective color vision , 2011, Vision Research.

[18]  D. Zaha,et al.  Significance of immunohistochemistry in breast cancer. , 2014, World journal of clinical oncology.

[19]  Wen-Hsiang Tsai,et al.  Moment-preserving thresolding: A new approach , 1985, Comput. Vis. Graph. Image Process..

[20]  Junnian Zheng,et al.  Ki67 is a promising molecular target in the diagnosis of cancer (review). , 2015, Molecular medicine reports.

[21]  Wen-Hsiang Tsai,et al.  Moment-preserving thresholding: a new approach , 1995 .

[22]  Shyang Chang,et al.  A new criterion for automatic multilevel thresholding , 1995, IEEE Trans. Image Process..

[23]  Michael Wagner,et al.  daime, a novel image analysis program for microbial ecology and biofilm research. , 2006, Environmental microbiology.

[24]  Anne E Carpenter,et al.  CellProfiler: image analysis software for identifying and quantifying cell phenotypes , 2006, Genome Biology.

[25]  Kevin W Eliceiri,et al.  NIH Image to ImageJ: 25 years of image analysis , 2012, Nature Methods.

[26]  Genee Y. Lee,et al.  Three-dimensional culture models of normal and malignant breast epithelial cells , 2007, Nature Methods.

[27]  Anne E Carpenter,et al.  A call for bioimaging software usability , 2012, Nature Methods.

[28]  N. Otsu.  A threshold selection method from gray level histograms , 1979 .

[29]  Marie-Liesse Asselin-Labat,et al.  The Ets transcription factor Elf5 specifies mammary alveolar cell fate. , 2008, Genes & development.

[30]  M L Mendelsohn,et al.  THE ANALYSIS OF CELL IMAGES * , 1966, Annals of the New York Academy of Sciences.

[31]  Nicolas Chenouard,et al.  Icy: an open bioimage informatics platform for extended reproducible research , 2012, Nature Methods.

[32]  M. Karnovsky,et al.  THF EARLY STAGES OF ABSORPTION OF INJECTED HORSERADISH PEROXIDASE IN THE PROXIMAL TUBULES OF MOUSE KIDNEY: ULTRASTRUCTURAL CYTOCHEMISTRY BY A NEW TECHNIQUE , 1966, The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society.

[33]  Nahed A. Soliman,et al.  Ki-67 as a prognostic marker according to breast cancer molecular subtype , 2016, Cancer biology & medicine.

[34]  Carolina Wählby,et al.  BlobFinder, a tool for fluorescence microscopy image cytometry , 2009, Comput. Methods Programs Biomed..

[35]  Mao-Jiun J. Wang,et al.  Image thresholding by minimizing the measures of fuzzines , 1995, Pattern Recognit..

[36]  F G Robertson,et al.  Loss of mammary epithelial prolactin receptor delays tumor formation by reducing cell proliferation in low-grade preinvasive lesions , 2007, Oncogene.

[37]  C. H. Li,et al.  An iterative algorithm for minimum cross entropy thresholding , 1998, Pattern Recognit. Lett..

[38]  Johannes E. Schindelin,et al.  Fiji: an open-source platform for biological-image analysis , 2012, Nature Methods.

[39]  A. Biankin,et al.  BAG-1 predicts patient outcome and tamoxifen responsiveness in ER-positive invasive ductal carcinoma of the breast , 2008, British Journal of Cancer.

[40]  Wassim Raffoul,et al.  A Preclinical Model for ERα-Positive Breast Cancer Points to the Epithelial Microenvironment as Determinant of Luminal Phenotype and Hormone Response. , 2016, Cancer cell.