This study successfully combined spatial omics analysis of overall chromatin accessibility and gene expression on the same tissue sample (spatial assay for transposase-accessible chromatin and RNA using sequencing (spatial ATAC–RNA-seq)), as well as performed simultaneous co-analysis of histone modifications and gene expression (spatial assay of cleavage under targets and tagmentation and RNA using sequencing (spatial CUT&Tag–RNA-seq)). To implement these approaches, the researchers introduced a two-dimensional grid of tissue pixels defined by spatial barcodes — two microfluidic chips placed sequentially in perpendicular directions on the tissue slice, introducing the spatial barcodes Ai (i = 1–50 or 100) and Bj (j = 1–50 or 100), respectively, to encode each pixel point. This system covers an area of about 16 square millimeters with near single-cell resolution; each pixel point is 20 micrometers and the total number of pixels encoded is up to 10,000. “Designing and optimizing the barcodes strategy is the most challenging part,” says Yanxiang Deng, a former postdoctoral researcher in the Fan laboratory and current assistant professor at the University of Pennsylvania. “We made a lot of efforts to ensure that the two omics assays can be compatible and addable, without interfering with each other.”
To further increase the precision of the spatial sequencing method, a combination with imaging technologies such as single-molecular fluorescence in situ hybridization (FISH) or single-molecule fluorescent imaging would be desirable. This would help to overcome a limitation of the method in which a single pixel may capture a half cell, one cell or, at times, several cells. Fan explains, “It’s not just a matter of adding one plus one to get two — this combined approach yields a much more enriched dataset.”
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