RT Journal Article
SR Electronic
T1 Workflow for high-dimensional flow cytometry analysis of T cells from tumor metastases
JF Life Science Alliance
JO Life Sci. Alliance
FD Life Science Alliance LLC
SP e202101316
DO 10.26508/lsa.202101316
VO 5
IS 10
A1 Cristina Faccani
A1 Gianluca Rotta
A1 Francesca Clemente
A1 Maya Fedeli
A1 Danilo Abbati
A1 Francesco Manfredi
A1 Alessia Potenza
A1 Achille Anselmo
A1 Federica Pedica
A1 Guido Fiorentini
A1 Chiara Villa
A1 Maria P Protti
A1 Claudio Doglioni
A1 Luca Aldrighetti
A1 Chiara Bonini
A1 Giulia Casorati
A1 Paolo Dellabona
A1 Claudia de Lalla
YR 2022
UL https://www.life-science-alliance.org/content/5/10/e202101316.abstract
AB We describe a multi-step high-dimensional (HD) flow cytometry workflow for the deep phenotypic characterization of T cells infiltrating metastatic tumor lesions in the liver, particularly derived from colorectal cancer (CRC-LM). First, we applied a novel flow cytometer setting approach based on single positive cells rather than fluorescent beads, resulting in optimal sensitivity when compared with previously published protocols. Second, we set up a 26-color based antibody panel designed to assess the functional state of both conventional T-cell subsets and unconventional invariant natural killer T, mucosal associated invariant T, and gamma delta T (γδT)-cell populations, which are abundant in the liver. Third, the dissociation of the CRC-LM samples was accurately tuned to preserve both the viability and antigenic integrity of the stained cells. This combined procedure permitted the optimal capturing of the phenotypic complexity of T cells infiltrating CRC-LM. Hence, this study provides a robust tool for high-dimensional flow cytometry analysis of complex T-cell populations, which could be adapted to characterize other relevant pathological tissues.