Ultrastructural Analysis of Trabecular Meshwork

Transmission electron microscopy (TEM) was used to evaluate the ultrastructure of the TM as described previously (Yun et al., 2014 (link)). After removing the iris, the limbus tissues (n = 3) from each group were ﬁxed in Karnovsky’s ﬁxative and divided into quarters of each tissue. Subsequently, the tissues were dehydrated and embedded in Epon and 65 nm Ultrathin sections were cut, stained with uranyl acetate (Electron Microscopy Sciences) and Reynold’s lead citrate (Fisher). Sections were photographed at 80 kV on a Jeol 1011 TEM for analysis. For evaluation of ER size, the boundary of ER on each TEM image was delineated and ER region was colored by photoshop (Adobe). Then, the area and perimeter of the ER was calculated by Image pro plus (Media Cybernetics). The ER size was displayed as ER area/ER perimeter (nm²/nm).

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Xiong S., Kumar A., Tian S., Taher E.E., Yang E., Kinchington P.R., Xia X, & Du Y. (2021). Stem cell transplantation rescued a primary open-angle glaucoma mouse model. eLife, 10, e63677.

Publication 2021

uranyl acetate Reynold’s lead citrate Jeol 1011 TEM Image pro plus

Citrate Electron microscopy Epon Iris Perimeter Tissues Transmission electron microscopy Uranyl acetate

Corresponding Organization :

Other organizations : Central South University, Xiangya Hospital Central South University, University of Pittsburgh, Research Institute of Ophthalmology, McGowan Institute for Regenerative Medicine

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

ER size (ER area/ER perimeter (nm^2/nm))

control variables

Iris removal
Limbus tissue samples (n = 3) from each group
Tissue fixation in Karnovsky's fixative
Tissue dehydration and embedding in Epon
Ultrathin sectioning at 65 nm
Staining with uranyl acetate and Reynold's lead citrate
Imaging on Jeol 1011 TEM at 80 kV

controls

Positive control: Not mentioned
Negative control: Not mentioned

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