Methods and compositions for labeling cells
A cell and labeling technology, which is used in the field of labeling cells and compositions, can solve the problems of inability to process multiple samples in parallel, limitations, etc.
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Embodiment 1
[0412] Example 1. Cells incubated with cholesterol-conjugated signature barcodes can be detected in sequencing libraries
[0413] Single cell sequencing libraries were prepared from cells incubated with and without cholesterol-conjugated signature barcodes and analyzed to assess the ability to detect signature barcodes in the processed libraries.
[0414] Briefly, cells were washed in culture medium after washing in PBS. Cells were counted and separated into 2 mL Eppendorf tubes and incubated for five minutes at room temperature with: (1) cholesterol-conjugated signature barcodes at a concentration of 1 uM; or (2) 1 uM signature barcodes only (i.e. , barcodes not conjugated to cholesterol moieties). After incubation, cells were washed three times in medium. Cells were then pooled and counted. The pooled population of cells is then dispensed into droplets as generally described elsewhere herein to produce droplets comprising: (1) a single cell; and (2) a releasable nucleic a...
Embodiment 2
[0417] Example 2. DNA Sequencing Results of Cholesterol Conjugated Signature Barcode Libraries
[0418]Jurkat cells were washed in medium after washing in PBS and then counted. 100,000 of these cells were split into 5 Eppendorf tubes (2 mL) to generate 5 different cell populations. Individual cell populations (four in total) were then incubated with 0.1 uM or 0.01 uM of cholesterol-conjugated signature barcodes (four in total, one per cell population) for five minutes at room temperature to generate "tagged" "one cell population with the first barcode (BC1), "tagged" one cell population with the second barcode (BC2), "tagged" one cell population with the third barcode (BC3), and "tagged" with the A cell population of the fourth barcode (BC4). One cell population was not incubated with cholesterol-conjugated signature barcodes (background population). The 5 cell populations were then washed in medium, pooled into individual tubes, and then counted to determine cell numbers. ...
Embodiment 3
[0426] Example 3. DNA sequencing results of antibody-conjugated signature barcode libraries
[0427] BioLegend "hashing" antibodies are provided that broadly target cell surface proteins in human cell types. Antibodies included a mixture of clones LNH94 (anti-CD298) and 2M2 (anti-[beta]2-microglobulin). Antibodies were pooled into different populations and barcoded with different characteristic barcodes. Jurkat, Raji and 293T cells were provided in separate populations and incubated with different antibody-associated signature barcodes. Jurkat cells were stained with antibodies barcoded with barcode #18 (BC18); Raji cells were stained with antibodies barcoded with barcode #19 (BC19); and 293T cells were stained with antibodies barcoded with barcode #20 (BC20). A total of 9,000 cells were loaded. The individual cell populations are then pooled. The pooled mixture is expected to include Jurkat cells comprising the signature barcode BC18, Raji cells comprising the signature b...
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