Cell　temperature　monitoring　is　of　great　importance　to　uncover　temperature-dependent　intracellular　events　and　regulate　cellular　functions.　However,　it　remains　a　great　challenge　to　precisely　probe　the　localized　temperature　status　in　living　cells.　Herein,　we　report　a　strategy　for　in　situ　temperature　mapping　on　an　immune　cell　membrane　for　the　first　time,　which　was　achieved　by　using　the　lanthanide-doped　upconversion　nanoparticles.　The　nanothermometer　was　designed　to　label　the　cell　membrane　by　combining　metabolic　labeling　and　click　chemistry　and　can　leverage　ratiometric　upconversion　luminescence　signals　to　in　situ　sensitively　monitor　temperature　variation　(1.4%　K-1).　Moreover,　a　purpose-built　upconversion　hyperspectral　microscope　was　utilized　to　synchronously　map　temperature　changes　on　T　cell　membrane　and　visualize　intracellular　Ca2+influx.　This　strategy　was　able　to　identify　a　suitable　temperature　status　for　facilitating　thermally　stimulated　calcium　influx　in　T　cells,　thus　enabling　high-efficiency　activation　of　immune　cells.　Such　findings　might　advance　understandings　on　thermally　dependent　biological　processes　and　their　regulation　methodology.　©　2022　American　Chemical　Society.　All　rights　reserved.