Framework for in vivo T cell screens.

Lauren E Milling,Samuel C Markson, Qin Tjokrosurjo, Nicole M Derosia, Ivy S L Streeter, Grant H Hickok, Ashlyn M Lemmen,Thao H Nguyen, Priyamvada Prathima,William Fithian,Marc A Schwartz,Nir Hacohen,John G Doench,Martin W LaFleur,Arlene H Sharpe

The Journal of experimental medicine(2024)

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Abstract
In vivo T cell screens are a powerful tool for elucidating complex mechanisms of immunity, yet there is a lack of consensus on the screen design parameters required for robust in vivo screens: gene library size, cell transfer quantity, and number of mice. Here, we describe the Framework for In vivo T cell Screens (FITS) to provide experimental and analytical guidelines to determine optimal parameters for diverse in vivo contexts. As a proof-of-concept, we used FITS to optimize the parameters for a CD8+ T cell screen in the B16-OVA tumor model. We also included unique molecular identifiers (UMIs) in our screens to (1) improve statistical power and (2) track T cell clonal dynamics for distinct gene knockouts (KOs) across multiple tissues. These findings provide an experimental and analytical framework for performing in vivo screens in immune cells and illustrate a case study for in vivo T cell screens with UMIs.
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