BibTex format
@article{Sparks:2025:pnasnexus/pgaf370,
author = {Sparks, H and Rowe-Brown, L and Alexandrov, Y and Gustafsson, N and Dvinskikh, L and Curry, N and Culley, J and Lee, M and Le, Marois A and Ratcliffe, CDH and Phillips, TA and Owczarek, C and Arias, Garcia M and Llanses, M and Suckert, T and Pelletier, J and Cortina, C and Hong, W and Garcia, E and Xu, Z and Zhang, S and Stassi, G and Batlle, E and Colombelli, J and Parsons, M and Bakal, C and Carragher, NO and Sahai, E and Dunsby, C},
doi = {pnasnexus/pgaf370},
journal = {PNAS Nexus},
title = {High content 3D imaging by dual-view oblique plane microscopy},
url = {http://dx.doi.org/10.1093/pnasnexus/pgaf370},
volume = {4},
year = {2025}
}
RIS format (EndNote, RefMan)
TY - JOUR
AB - Oblique plane microscopy (OPM) is a form of light-sheet fluorescence microscopy (LSFM) employing a single microscope objective at the sample for both fluorescence excitation and detection. Dual-view OPM (dOPM) is an optically folded form of OPM. We present an improved dOPM system employing a 60×/1.2NA water immersion primary objective and measure the spatial resolution and fluorescence collection efficiency for illumination angles of 35° and 45° with respect to the coverslip. Illumination at 35° provides slightly better lateral resolution and collection efficiency. Collection efficiency measurements are compared to a full vectorial raytracing simulation of the system. Using a light-sheet angle of 35°, the median bead FWHM for 100nm diameter fluorescent beads in x, y, and z and the optical sectioning strength were measured over a volume of 100 × 100 × 100μm3 to be 0.29, 0.31, 0.83, and 2.45–3.00μm, respectively when the two dOPM views are fused. We demonstrate less photobleaching in time-lapse dOPM of live mEmerald-expressing organoids compared to widefield epi-fluorescence z-stack imaging under the condition of equal detected fluorescence signal from a point object in focus. We demonstrate dOPM for multifield-of-view 3D imaging of biological samples in 96-well plates and apply it to imaging cells in collagen gel and quantifying the FUCCI cell-cycle reporter to provide drug dose–response curves in spheroids. We also use it to perform time-lapse multifield-of-view imaging and demonstrate the detection of organoid lumen closure and reopening, organoid migration within a collagen gel and observing dynamic events in arrays of ex vivo tissue slices.
AU - Sparks,H
AU - Rowe-Brown,L
AU - Alexandrov,Y
AU - Gustafsson,N
AU - Dvinskikh,L
AU - Curry,N
AU - Culley,J
AU - Lee,M
AU - Le,Marois A
AU - Ratcliffe,CDH
AU - Phillips,TA
AU - Owczarek,C
AU - Arias,Garcia M
AU - Llanses,M
AU - Suckert,T
AU - Pelletier,J
AU - Cortina,C
AU - Hong,W
AU - Garcia,E
AU - Xu,Z
AU - Zhang,S
AU - Stassi,G
AU - Batlle,E
AU - Colombelli,J
AU - Parsons,M
AU - Bakal,C
AU - Carragher,NO
AU - Sahai,E
AU - Dunsby,C
DO - pnasnexus/pgaf370
PY - 2025///
SN - 2752-6542
TI - High content 3D imaging by dual-view oblique plane microscopy
T2 - PNAS Nexus
UR - http://dx.doi.org/10.1093/pnasnexus/pgaf370
UR - https://academic.oup.com/pnasnexus/article/4/12/pgaf370/8342766
VL - 4
ER -