title: Application of multispectral imaging in quantitative immunohistochemistry study of breast cancer: a comparative study.
abstract: Multispectral imaging (MSI) based on imaging and spectroscopy, as relatively novel to the field of histopathology, has been used in biomedical multidisciplinary researches. We analyzed and compared the utility of multispectral (MS) versus conventional red-green-blue (RGB) images for immunohistochemistry (IHC) staining to explore the advantages of MSI in clinical-pathological diagnosis. The MS images acquired of IHC-stained membranous marker human epidermal growth factor receptor 2 (HER2), cytoplasmic marker cytokeratin5/6 (CK5/6), and nuclear marker estrogen receptor (ER) have higher resolution, stronger contrast, and more accurate segmentation than the RGB images. The total signal optical density (OD) values for each biomarker were higher in MS images than in RGB images (all P < 0.05). Moreover, receiver operator characteristic (ROC) analysis revealed that a greater area under the curve (AUC), higher sensitivity, and specificity in evaluation of HER2 gene were achieved by MS images (AUC = 0.91, 89.1 %, 83.2 %) than RGB images (AUC = 0.87, 84.5, and 81.8 %). There was no significant difference between quantitative results of RGB images and clinico-pathological characteristics (P > 0.05). However, by quantifying MS images, the total signal OD values of HER2 positive expression were correlated with lymph node status and histological grades (P = 0.02 and 0.04). Additionally, the consistency test results indicated the inter-observer agreement was more robust in MS images for HER2 (inter-class correlation coefficient (ICC) = 0.95, r s = 0.94), CK5/6 (ICC = 0.90, r s = 0.88), and ER (ICC = 0.94, r s = 0.94) (all P < 0.001) than that in RGB images for HER2 (ICC = 0.91, r s = 0.89), CK5/6 (ICC = 0.85, r s = 0.84), and ER (ICC = 0.90, r s = 0.89) (all P < 0.001). Our results suggest that the application of MS images in quantitative IHC analysis could obtain higher accuracy, reliability, and more information of protein expression in relation to clinico-pathological characteristics versus conventional RGB images. It may become an optimal IHC digital imaging system used in quantitative pathology.
authors.affiliation: Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors & Hubei Cancer Clinical Study Center, Wuhan, 430071, China.
authors.forename: Wen-Lou
authors.initials: WL
authors.lastname: Liu
authors.affiliation: Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors & Hubei Cancer Clinical Study Center, Wuhan, 430071, China.
authors.forename: Lin-Wei
authors.initials: LW
authors.lastname: Wang
authors.affiliation: Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors & Hubei Cancer Clinical Study Center, Wuhan, 430071, China.
authors.forename: Jia-Mei
authors.initials: JM
authors.lastname: Chen
authors.affiliation: Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors & Hubei Cancer Clinical Study Center, Wuhan, 430071, China.
authors.forename: Jing-Ping
authors.initials: JP
authors.lastname: Yuan
authors.affiliation: Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors & Hubei Cancer Clinical Study Center, Wuhan, 430071, China.
authors.forename: Qing-Ming
authors.initials: QM
authors.lastname: Xiang
authors.affiliation: Department of Pathology, Zhongnan Hospital of Wuhan University, Wuhan, 430071, China.
authors.forename: Gui-Fang
authors.initials: GF
authors.lastname: Yang
authors.affiliation: Key State Laboratory of Software Engineering, School of Computer, Wuhan University, Wuhan, 430072, China.
authors.forename: Ai-Ping
authors.initials: AP
authors.lastname: Qu
authors.affiliation: Key State Laboratory of Software Engineering, School of Computer, Wuhan University, Wuhan, 430072, China.
authors.forename: Juan
authors.initials: J
authors.lastname: Liu
authors.affiliation: Department of Oncology, Zhongnan Hospital of Wuhan University, Hubei Key Laboratory of Tumor Biological Behaviors & Hubei Cancer Clinical Study Center, Wuhan, 430071, China. liyansd2@163.com.|Department of Peritoneal Cancer Surgery, Beijing Shijitan Hospital of Capital Medical University, Beijing, 100038, China. liyansd2@163.com.|Department of Oncology, Zhongnan Hospital of Wuhan University; Department of Peritoneal Cancer Surgery, Beijing Shijitan Hospital of Capital Medical University, No 10, Tieyi Road, Yangfangdian, Haidian District, Beijing, 100038, China. liyansd2@163.com.
authors.forename: Yan
authors.initials: Y
authors.lastname: Li
mesh_terms: D000328:Adult; D000368:Aged; D014408:Biomarkers, Tumor / Q000096:biosynthesis* / Q000302:isolation & purification; D001943:Breast Neoplasms / Q000000981:diagnostic imaging* / Q000235:genetics / Q000473:pathology; D047628:Estrogen Receptor alpha / Q000096:biosynthesis* / Q000302:isolation & purification; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D053555:Keratin-5 / Q000096:biosynthesis* / Q000302:isolation & purification; D008875:Middle Aged; D057054:Molecular Imaging / Q000379:methods; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000096:biosynthesis* / Q000302:isolation & purification
chemical_list: D014408:Biomarkers, Tumor; D047628:Estrogen Receptor alpha; D053555:Keratin-5; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
keywords: Immunohistochemistry; Invasive breast cancer; Multispectral imaging; Optical density; Quantitative image analysis