Modal focal adaptive optics for Bessel-focus two-photon fluorescence microscopy

Adaptive optics (AO) significantly enhances microscopic imaging by correcting optical aberrations. However, its application in Bessel-focus two-photon fluorescence microscopy (2PFM) has been underexplored. Here we introduce a novel modal focal AO (MFAO) method for Bessel-focus 2PFM, utilizing Zernike annular modes and a focal AO technique that enable aberration measurement, correction, and Bessel-focus generation with a single spatial light modulator (SLM). This approach offers a simpler and more cost-effective alternative to a previous zonal focal AO method, achieving comparable performance in Bessel-focus 2PFM. We demonstrate MFAO's efficacy in correcting aberrations through imaging of fixed mouse brain slices and in vivo zebrafish larvae and mouse brains, advancing AO applications in non-traditional microscopy with non-circular pupils.