TY  - JOUR
AU  - Wellach, Kathrin
AU  - Riemer, Angelika
TI  - Highly sensitive live-cell imaging-based cytotoxicity assay enables functional validation of rare epitope-specific CTLs.
JO  - Frontiers in immunology
VL  - 16
SN  - 1664-3224
CY  - Lausanne
PB  - Frontiers Media
M1  - DKFZ-2025-01088
SP  - 1558620
PY  - 2025
N1  - #EA:D410#LA:D410#
AB  - Many immunotherapeutic approaches aim to induce epitope-specific T-cell cytotoxicity. However, the identification-and especially the functional validation-of suitable epitopes by in vitro cytotoxicity assays can be challenging, particularly when the number of available epitope-specific cytotoxic T cells (CTLs) is limited. Here, we present a highly sensitive image-based cytotoxicity assay that allows the functional analysis of rare epitope-specific T cells. The live-cell imaging-based setup combines transient red labeling of target cells with a green caspase 3/7 probe, allowing reliable measurement of the fraction of apoptotic target cells. Time-course analysis enables the monitoring of subtle differences. This highly flexible assay can be applied to assess the killing of either target cells with endogenous epitope presentation or those artificially loaded with the epitope of interest. Analysis of assay sensitivity demonstrated that cytotoxicity mediated by as few as 0.1
KW  - Humans
KW  - T-Lymphocytes, Cytotoxic: immunology
KW  - T-Lymphocytes, Cytotoxic: metabolism
KW  - Epitopes, T-Lymphocyte: immunology
KW  - Cytotoxicity, Immunologic
KW  - Lymphocyte Activation Gene 3 Protein
KW  - Cytotoxicity Tests, Immunologic: methods
KW  - Cell Line, Tumor
KW  - Interferon-gamma: metabolism
KW  - Programmed Cell Death 1 Receptor: metabolism
KW  - Programmed Cell Death 1 Receptor: immunology
KW  - Apoptosis
KW  - Caspase 3: metabolism
KW  - T cells (Other)
KW  - cytotoxicity (Other)
KW  - epitopes (Other)
KW  - live-cell imaging (Other)
KW  - rare CTL populations (Other)
KW  - Epitopes, T-Lymphocyte (NLM Chemicals)
KW  - Lymphocyte Activation Gene 3 Protein (NLM Chemicals)
KW  - Interferon-gamma (NLM Chemicals)
KW  - Lag3 protein, human (NLM Chemicals)
KW  - Programmed Cell Death 1 Receptor (NLM Chemicals)
KW  - Caspase 3 (NLM Chemicals)
LB  - PUB:(DE-HGF)16
C6  - pmid:40406125
C2  - pmc:PMC12095279
DO  - DOI:10.3389/fimmu.2025.1558620
UR  - https://inrepo02.dkfz.de/record/301580
ER  -