Osmoregulation is critical for cell and tissue survival yet there are relatively few methods available to determine osmotic gradients from water and elemental concentration either in single cells or across multiple cellular layers of tissue. X-ray microanalysis of frozen-hydrated preparations in a scanning electron microscope is one such powerful, sensitive, nondestructive technique. Here we use x-ray microanalysis to quantitatively analyse intracellular element concentrations and oxygen concentrations, as a proxy for water concentrations, in selected individual cells of the posterior eye. Using frozen-hydrated preparations of the retinal complex of chicken eyes, it is shown that structural preservation is sufficient to identify cell layers and individual cells. The quantitative analysis of selected areas in the photoreceptor layer, inner nuclear layer and ganglion cell layer, where specific cell types were known to be present, provided measurements of intracellular element concentrations comparable with the analysis of individual cells. It is also shown that in the cells of the retinal pigment epithelium and outer photoreceptor segments elemental analyses were reasonably consistent at the cellular level in different depth levels of the same sample. Comparison of oxygen concentrations, as a proxy for water concentration, at two accelerating voltages (15 and 5 kV) indicated that at 15 kV oxygen concentration was largely derived from intracellular water. Water concentrations could be calculated and concentrations of diffusible elements (Na, K) could be defined in mmol/L. From the latter it is possible to calculate osmotic concentrations of individual cells and osmotic gradients across the tissue.

中文翻译：

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一种用于测量后眼细胞和组织中与渗透压调节有关的体内元素和水浓度的 X 射线微量分析方法

渗透压调节对于细胞和组织的存活至关重要，但可用于从水和元素浓度确定渗透梯度的方法相对较少，无论是在单个细胞中还是在组织的多个细胞层中。在扫描电子显微镜中对冷冻水合制剂进行 X 射线微量分析是一种强大、灵敏、无损的技术。在这里，我们使用 X 射线微量分析来定量分析后眼选定单个细胞中的细胞内元素浓度和氧浓度，作为水浓度的代表。使用鸡眼视网膜复合体的冷冻水合制剂，表明结构保存足以识别细胞层和单个细胞。感光层中选定区域的定量分析，已知存在特定细胞类型的内核层和神经节细胞层提供了与单个细胞分析相当的细胞内元素浓度测量值。它还表明，在视网膜色素上皮细胞和外部感光器节段的细胞中，元素分析在同一样品的不同深度水平的细胞水平上是相当一致的。在两个加速电压（15 和 5 kV）下，作为水浓度代表的氧浓度的比较表明，在 15 kV 下，氧浓度主要来自细胞内水。可以计算水浓度，并且可以以 mmol/L 定义可扩散元素（Na、K）的浓度。