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Updated: May 19, 2026
Quantitation of Protein Expression and Co-localization Using Multiplexed Immuno-histochemical Staining and Multispectral Imaging
Published on: April 8, 2016
A colorful future of quantitative pathology: validation of Vectra technology using chromogenic multiplexed
Wei Huang1, Kenneth Hennrick, Sally Drew
1Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, Madison, WI 53792, USA. whuang23@wisc.edu
Abstract:
The Vectra platform (Caliper Life Sciences, Hopkinton, MA) is an advanced multispectral imaging system for biomarker quantitation in tissue microarray or intact tissue sections. This is the first study to validate its reliability for quantitating spatially overlapping biomarkers using chromogenic multiplexed immunohistochemistry on prostate tissue microarrays. Two tissue microarray cohorts (an outcome tissue microarray and a progression tissue microarray) were used. The outcome tissue microarray cohort consists of 462 duplicate cores with more than 5-year outcome information. The progression tissue microarray cohort consists of 384 duplicate cores from different disease (stage) groups. The tissue microarray slides were stained with different combinations of antibodies (anti-androgen receptor, anti-E-cadherin, anti-erythroblastosis virus E26 oncogene-related gene product, and anti-α-methylacyl-CoA racemase). Three outcome tissue microarrays were stained with androgen receptor + erythroblastosis virus E26 oncogene-related gene + E-cadherin (outcome tissue microarray 1), androgen receptor + E-cadherin (outcome tissue microarray 2), and erythroblastosis virus E26 oncogene-related gene + E-cadherin (outcome tissue microarray 3), respectively. One progression tissue microarray section was stained with E-cadherin and α-methylacyl-CoA racemase; tissue microarray slides were then scanned with the Vectra platform. Biomarker expression analysis was performed with Vectra software-Nuance 3.0.0, and inForm 1.2. IBM SPSS Statistics 19 was used for statistical and correlation analysis (SPSS, Chicago, IL). Close concordance was found between the triple- and double-immunostaining assays used for quantitating spatially overlapping biomarkers androgen receptor and erythroblastosis virus E26 oncogene-related gene using outcome tissue microarrays (r = 0.897 for androgen receptor and 0.613 for erythroblastosis virus E26 oncogene-related gene, respectively). α-Methylacyl-CoA racemase and E-cadherin expression levels measured in progression tissue microarray were consistent with previously published data by other groups. In conclusion, Vectra technology is reliable for objective and high-throughput biomarker quantitation and colocalization study using chromogenic multiplexed immunohistochemistry.
