TY - JOUR
T1 - Three-dimensional deconvolution processing for STEM cryotomography
AU - Waugh, Barnali
AU - Wolf, Sharon G.
AU - Fass, Deborah
AU - Branlund, Eric
AU - Kam, Zvi
AU - Sedat, John W.
AU - Elbaum, Michael
N1 - This record is sourced from MEDLINE/PubMed, a database of the U.S. National Library of Medicine. We are grateful to Muthuvel Arigovindan, Shahar Seifer, Charles Kervrann, David Agard, and David DeRosier for enlightening discussions. This work was supported in part by a grant from the Israel Science Foundation. M.E. is the incumbent of the Sam and Ayala Zacks Professorial Chair and head of the Irving and Cherna Moskowitz Center for Nano and Bio-Nano Imaging. The laboratory of M.E. has benefitted from the historical generosity of the Harold Perlman family. Author contributions: J.W.S. and M.E. designed research; B.W., S.G.W., and D.F. performed research; E.B. and M.E. contributed new reagents/analytic tools; B.W., Z.K., J.W.S., and M.E. analyzed data; and M.E. wrote the paper.
PY - 2020/11/3
Y1 - 2020/11/3
N2 - The complex environment of biological cells and tissues has motivated development of three-dimensional (3D) imaging in both light and electron microscopies. To this end, one of the primary tools in fluorescence microscopy is that of computational deconvolution. Wide-field fluorescence images are often corrupted by haze due to out-of-focus light, i.e., to cross-talk between different object planes as represented in the 3D image. Using prior understanding of the image formation mechanism, it is possible to suppress the cross-talk and reassign the unfocused light to its proper source post facto. Electron tomography based on tilted projections also exhibits a cross-talk between distant planes due to the discrete angular sampling and limited tilt range. By use of a suitably synthesized 3D point spread function, we show here that deconvolution leads to similar improvements in volume data reconstructed from cryoscanning transmission electron tomography (CSTET), namely a dramatic in-plane noise reduction and improved representation of features in the axial dimension. Contrast enhancement is demonstrated first with colloidal gold particles and then in representative cryotomograms of intact cells. Deconvolution of CSTET data collected from the periphery of an intact nucleus revealed partially condensed, extended structures in interphase chromatin.
AB - The complex environment of biological cells and tissues has motivated development of three-dimensional (3D) imaging in both light and electron microscopies. To this end, one of the primary tools in fluorescence microscopy is that of computational deconvolution. Wide-field fluorescence images are often corrupted by haze due to out-of-focus light, i.e., to cross-talk between different object planes as represented in the 3D image. Using prior understanding of the image formation mechanism, it is possible to suppress the cross-talk and reassign the unfocused light to its proper source post facto. Electron tomography based on tilted projections also exhibits a cross-talk between distant planes due to the discrete angular sampling and limited tilt range. By use of a suitably synthesized 3D point spread function, we show here that deconvolution leads to similar improvements in volume data reconstructed from cryoscanning transmission electron tomography (CSTET), namely a dramatic in-plane noise reduction and improved representation of features in the axial dimension. Contrast enhancement is demonstrated first with colloidal gold particles and then in representative cryotomograms of intact cells. Deconvolution of CSTET data collected from the periphery of an intact nucleus revealed partially condensed, extended structures in interphase chromatin.
U2 - 10.1073/pnas.2000700117
DO - 10.1073/pnas.2000700117
M3 - مقالة
C2 - 33077585
SN - 0027-8424
VL - 117
SP - 27374
EP - 27380
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 44
ER -