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Clarification and confocal imaging of the nonhuman primate placental micro-anatomy
摘要： Geometry of the placental villous vasculature is a key determinant of maternal–fetal nutrient exchange for optimal fetal growth. Recent advances in tissue clarification techniques allow for deep high-resolution imaging with confocal microscopy; however, the methodology lacks a signal:noise ratio of sufficient magnitude to allow for quantitative analysis. Thus, we sought to develop a reproducible method to investigate the 3D vasculature of the nonhuman primate placenta for subsequent data analysis. Fresh placental tissue was dissected, formalin fixed, clarified using a modified Visikol? protocol and immunolabeled for CD31 (fetal endothelium) and cytokeratin-7 (villous trophoblast) for confocal imaging of the microanatomy. We present a detailed clarification and staining protocol augmented for imaging of nonhuman primate placental tissue. The image stacks generated by this refined staining method and our data acquisition parameters can be analyzed quantitatively to provide insights regarding the villous and vascular micro-anatomy of the placenta.
关键词： placenta,immunohistochemistry,tissue clarification,nonhuman primate,confocal microscopy
更新于2025-09-10 09:29:36
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Total Retinal Blood Flow in a Nonhuman Primate Optic Nerve Transection Model Using Dual-Beam Bidirectional Doppler FD-OCT and Microsphere Method
摘要： PURPOSE. We validated noninvasive Doppler-optical coherence tomography (OCT) blood ?ow measurements against the terminal microsphere method in a surgical induced optic nerve transection nonhuman primate model. METHODS. In 6 nonhuman primates, total retinal blood ?ow (TRBF) was measured with a custom-built dual-beam bidirectional Doppler Fourier Domain (FD)-OCT. Peripapillary retinal nerve ?ber layer thickness (RNFLT) was measured by Spectralis spectral-domain (SD)-OCT. Measurements were performed every 10 to 15 days before and after unilateral optic nerve transection (ONT) until RNFLT was reduced by more than 40% from baseline. Before the animals were killed, TRBF was measured using the microsphere technique. RESULTS. A signi?cant correlation between all arterial and venous Doppler OCT TRBF measurements was found in ONT and contralateral control eyes (both P < 0.01, n ? 6). The Bland-Altman analysis showed a bias of 0.57 in the ONT group and 0.02 in the contralateral control group. Also, excellent agreement was observed between Doppler OCT and microsphere measurements (P < 0.01, r ? 0.976, bias ? 0.54). After ONT, TRBF and RNFLT decreased by (cid:2)51% 6 42% and (cid:2)44% 6 2% (n ? 5), respectively. In the contralateral control eyes, TRBF and RNFLT were unchanged. CONCLUSIONS. Very good accordance was found between TRBF measurements, obtained with dual-beam bidirectional Doppler FD-OCT and the microsphere method. It also was possible to monitor changes over time in TRBF after ONT with Doppler OCT. These ?ndings highlight the accuracy and potential of noninvasive Doppler OCT to provide valuable information for detecting early changes in ocular disease in future.
关键词： microspheres,optic nerve transection,total retinal blood ?ow,Doppler OCT,nonhuman primate
更新于2025-09-09 09:28:46