Single Cell Reactomics: Real‐Time Single‐Cell Activation Kinetics of Optically Trapped Macrophages

Abstract

Macrophages are well known for their role in immune responses and tissue homeostasis. They can polarize towards various phenotypes in response to biophysical and biochemical stimuli. However, little is known about the early kinetics of macrophage polarization in response to single biophysical or biochemical stimuli. Our approach, combining optical tweezers, confocal fluorescence microscopy, and microfluidics, allows us to isolate single macrophages and follow their immediate responses to a biochemical stimulus in real‐time. This strategy enables live‐cell imaging at high spatiotemporal resolution and omits surface adhesion and cell–cell contact as biophysical stimuli. The approach is validated by successfully following the early phase of an oxidative stress response of macrophages upon phorbol 12‐myristate 13‐acetate (PMA) stimulation, allowing detailed analysis of the initial macrophage response upon a single biochemical stimulus within seconds after its application, thereby eliminating delay times introduced by other techniques during the stimulation procedure. Hence, an unprecedented view of the early kinetics of macrophage polarization is provided.

In Situ Monitoring of Hydrogen Peroxide Released from Living Cells Using a ZIF-8-Based Surface-Enhanced Raman Scattering Sensor

“…Typically, PMA-activated protein kinase C triggered the oxidation of oxygen by NOX (NADPH oxidase), along with the formation of • O 2 – . Under the reduction effect of superoxide dismutase (SOD1), H 2 O 2 generated and released to the extracellular environment to initiate intracellular signaling. , The monitoring of ROS released from cells under PMA stimulation was performed by the AuNPs@2-MHQ@ZIF-8 SERS sensor (Figure B). Figure C shows a relatively high H 2 O 2 level with the addition of 3 μg/mL PMA concentration.…”

Biophysical Approaches for Applying and Measuring Biological Forces

“…Recently, OTs have also been combined with other instruments, such as fluorescence microscopy and microfluidic platforms, to simultaneously detect multiple signals. [ 153 ] In the cross‐instrument approach developed by Vasse et al, a dual‐beam optical trap is employed to capture an isolated single macrophage, hence greatly reducing the effect caused by cell‐cell interactions and surface adhesion. [ 153a ] Their strategy allows the detection and tracking of the response of a cell to single biomechanical or biochemical stimuli at high spatiotemporal resolution in real time.…”

“…[ 153 ] In the cross‐instrument approach developed by Vasse et al, a dual‐beam optical trap is employed to capture an isolated single macrophage, hence greatly reducing the effect caused by cell‐cell interactions and surface adhesion. [ 153a ] Their strategy allows the detection and tracking of the response of a cell to single biomechanical or biochemical stimuli at high spatiotemporal resolution in real time. Also, optical traps offer a powerful platform for single molecular‐ or single‐cell‐level microrheology measurements and have been successfully exploited for a variety of applications.…”

Non-Invasive Dynamic Reperfusion of Microvessels In Vivo Controlled by Optical Tweezers

“…In this context, optical tweezers ( Ashkin et al, 1986 ; Neuman and Block, 2004 ) have been used to manipulate and investigate microscopic particles for many years, and the ability to study living cells at the single-cell level have been proven ( Zhang and Liu, 2008 ; Zhao et al, 2020 ; Vasse et al, 2021 ). Significant progress has been made in the manipulation of biological cells rotation with optical tweezers for orientation-based cell surgery ( Xie et al, 2015a ; Xie et al, 2015b ; Xie et al, 2018a ).…”