This study introduces two novel red-emissive BODIPY dyes, each functionalized with methoxystyryl groups at the 3- and 5-positions to bathochromically shift spectral bands. Obtained via facile synthetic pathways, the dyes can be excited using red single-photon or near-infrared two-photon excitation. The latter is of key importance, with long-wavelength excitation being advantageous for live-cell microscopy as it offers increased tissue penetration, reduced scattering, and minimal autofluorescence.Comprehensive structural, computational, and photophysical characterisation highlight the suitability of both dyes for use in live-cell laser scanning confocal microscopy (LSCM) of mouse embryonic fibroblast (NIH-3T3) cells. The materials are characterised by high brightness with high molar absorption coefficients and photoluminescence quantum yields. Additionally, two-photon absorption cross-sections were measured as 723 GM and 502 GM (1 GM = 10–50 cm4 s photon−1) at 840 nm, respectively, making the dyes suitable for multiphoton microscopy applications.Dosing protocols were developed for single-photon and multiphoton LSCM, and co-staining confirmed strong localisation within lipid droplets (LDs). LDs are an important organelle to study regarding manifestation of diseases such as diabetes and cancer, as well as monitoring of distribution of lipophilic intracellular material. Cytotoxicity assays at a range of concentrations (500 nM – 5 μM) for up to 24 h revealed both dyes can be used for prolonged live-cell microscopy without significant alterations to cellular viability
