Cell hashing vs barcoding
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7 weeks ago
Cheng Wei • 0

Hi everyone,

I cant seem to unable to differentiate between cell hashing , for example https://cite-seq.com/cell-hashing/, or cellplex(by 10x) with cell barcoding (used by 10x) for example. What is it with cell hashing that makes it different?

barcoding hashing • 296 views
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7 weeks ago
ATpoint 54k

Cell barcoding means that each single cell in an oil droplet will be equipped with a specific DNA sequence that is incorporated into the final library fragments. Read1 if the paired-end sequencing will read this Cellular Barcode (CB). Each DNA sequencing fragment that has been produced inside the same oil droplet (and by this ideally from the same single cell unless a doublet has been captured) has now the same CB. It is a connector between a cell and all the reads produced from DNA fragments that come from the original cDNA of that cell.

Cell hashing is basically "labelling" a group of cells with a specific antibody that carries a certain sequence. Imagine you isolate from a single mouse cells from heart, lung and spleen. You do this separately, get 1000 cells each after some FACS sorting but want to pool these cells before doing the 10X library preparation. Obviously you want to know which cells did originally belong to which organ. This is where hasing comes into play. Before pooling the cells you add an antibody to each tube with cells that binds to a uniquitous protein on the surface of the cell. The antibody is now attached to the cell surface, the cells are pooled and submitted to the emulsification into the droplets and all subsequent library preparation steps. The DNA sequence attached to the antibody is (like the cDNA that is created) incorporated into the final sequencing library and by this ends up as reads in the fastq files. This is a connector between a predefined group of cells and all reads so you can later easily tell which cells did originally come from the same pool of cells.

Summary: CBs connect each individual cells with the reads that belong to the cDNA of that cell. Hashing allows researchers to pool a predefined selection of cells in the same library preparation procedure while still being able to later distinguish the pools.

Does that make sense to you?

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Yep, it makes sense to me. Does this mean that cell barcoding is something we do on top of barcoding to differentiate the cells?

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Hashing is optional, barcoding is not.

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