PT  - JOURNAL ARTICLE
AU  - Yury Goltsev
AU  - Nikolay Samusik
AU  - Julia Kennedy-Darling
AU  - Salil Bhate
AU  - Matthew Hale
AU  - Gustavo Vasquez
AU  - Garry P. Nolan
TI  - Deep profiling of mouse splenic architecture with CODEX multiplexed imaging
AID  - 10.1101/203166
DP  - 2017 Jan 01
TA  - bioRxiv
PG  - 203166
4099  - http://biorxiv.org/content/early/2017/10/20/203166.short
4100  - http://biorxiv.org/content/early/2017/10/20/203166.full
AB  - A cytometric imaging approach, called CO-Detection by indEXing (CODEX), that enables high parameter multiplexing of antibody-tagged target epitopes is used here to create high parameter imaging datasets of normal mouse and lupus (MRL/lpr) spleens. In this procedure, antibody binding events are rendered iteratively using DNA barcodes, fluorescent dNTP analogs, and an in-situ polymerization-based indexing procedure. Fluorescent signals from multiple rounds of indexing are computationally combined into a multi-channel image stack and subjected to image segmentation and quantification. A segmentation and linear model algorithm was developed to accurately quantify membrane antigen levels on dissociated cells as well as tissue sections. Leveraging the spatially resolved nature of CODEX multiplexed single-cell imaging data, quantitative de novo characterization of lymphoid tissue architecture was enabled and overlaid onto previously described morphological features. We observed an unexpected, profound impact of the cellular neighborhood on the expression of protein receptors on immune cells. By comparing normal murine spleen to spleens from animals with systemic autoimmune disease (MRL/lpr), extensive and previously uncharacterized splenic cell interaction dynamics in the healthy versus diseased state was observed. The fidelity of multiplexed imaging data analysis demonstrated here will allow deep proteomic analysis and systematic characterization of complex tissue architecture in normal and clinically aberrant samples.