Long-term nondestructive monitoring of cells is of significant importance for understanding cell proliferation, cell signaling, cell death, and other processes. However, traditional monitoring methods are limited to a certain range of testing conditions and may reduce cell viability. Here, we present a microgap, multishot electroporation (M2E) system for monitoring cell recovery for up to ∼2 h using ∼5 V pulses and with excellent cell viability using a medium cell population. Electric field simulations reveal the bias-voltage- and gap-size-dependent electric field intensities in the M2E system. In addition to excellent transparency with low cell toxicity, the M2E system does not require specialized components, expensive materials, complicated fabrication processes, or cell manipulations; it just consists of a micrometer-sized pattern and a low-voltage square-wave generator. Ultimately, the M2E system can offer a long-term and nontoxic method of cell monitoring.