TY - CHAP
T1 - Quantitation of IRF3 nuclear translocation in heterogeneous cellular populations from cervical tissue using imaging flow cytometry
AU - Trifonova, Radiana T.
AU - Barteneva, Natalie
PY - 2018/1/1
Y1 - 2018/1/1
N2 - Imaging flow cytometry (IFC) has become a powerful tool for studying the activation of transcriptional factors in heterogeneous cell populations in high-content imaging mode. With considerable interest to the clinical development of IFC, the question becomes how we can accelerate its application to solid tissues. We developed the first IFC-based procedure to quantify the nuclear translocation of interferon regulatory factor (IRF) 3, an important measure of induction of type I interferon antiviral response, in primary human immune cells including in solid tissues. After tissue digestion and protocol optimization by spectral flow cytometry, cell suspension is stained for intracellular IRF3 and acquired by IFC. Image analysis is performed using an optimized nuclear mask and similarity score parameter to correlate the location of IRF3 staining and a nuclear dye. The technique measures IRF3 activation at a single cell level and can detect small changes in the percent of activated cells providing objective quantitative data for statistical analysis.
AB - Imaging flow cytometry (IFC) has become a powerful tool for studying the activation of transcriptional factors in heterogeneous cell populations in high-content imaging mode. With considerable interest to the clinical development of IFC, the question becomes how we can accelerate its application to solid tissues. We developed the first IFC-based procedure to quantify the nuclear translocation of interferon regulatory factor (IRF) 3, an important measure of induction of type I interferon antiviral response, in primary human immune cells including in solid tissues. After tissue digestion and protocol optimization by spectral flow cytometry, cell suspension is stained for intracellular IRF3 and acquired by IFC. Image analysis is performed using an optimized nuclear mask and similarity score parameter to correlate the location of IRF3 staining and a nuclear dye. The technique measures IRF3 activation at a single cell level and can detect small changes in the percent of activated cells providing objective quantitative data for statistical analysis.
KW - Cellular heterogeneity
KW - Imaging flow cytometry
KW - IRF3
KW - Nucleocytoplasmic translocation
KW - Signal transduction
KW - Solid tissue
UR - http://www.scopus.com/inward/record.url?scp=85042559438&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85042559438&partnerID=8YFLogxK
U2 - 10.1007/978-1-4939-7680-5_8
DO - 10.1007/978-1-4939-7680-5_8
M3 - Chapter
AN - SCOPUS:85042559438
T3 - Methods in Molecular Biology
SP - 125
EP - 153
BT - Methods in Molecular Biology
PB - Humana Press Inc.
ER -