Abstract：
The polysaccharide capsule of Cryptococcus neoformans is the primary virulence factor and one of the most commonly studied aspects of this pathogenic yeast. Capsule size varies widely between strains, has the ability to grow rapidly when introduced to stressful or low-nutrient conditions, and has been positively correlated with strain virulence. For these reasons, the size of the capsule is of great interest to C. neoformans researchers. Inducing the growth of the C. neoformans capsule is used during phenotypic testing to help understand the effects of different treatments on the yeast or size differences between strains. Here, we describe one of the standard methods of capsule induction and detail two accepted methods of staining: (i) India ink, a negative stain, used in conjunction with conventional light microscopy and (ii) co-staining with fluorescent dyes of both the cell wall and capsule followed by confocal microscopy. Finally, we outline how to measure capsule diameter manually and offer a protocol for automated diameter measurement of India ink-stained samples using computational image analysis.
摘要：
新生隐球菌的多糖胶囊是主要的毒力因子，也是这种致病性酵母最常研究的方面之一。胶囊大小在菌株之间差异很大，当被引入压力或低营养条件时，具有快速生长的能力，与毒株毒力呈正相关。由于这些原因,胶囊的大小是C.新生科学家非常感兴趣的。在表型测试过程中使用诱导新生梭菌胶囊的生长来帮助理解不同处理对酵母的影响或菌株之间的大小差异。这里,我们描述了胶囊诱导的标准方法之一，并详细介绍了两种公认的染色方法：（i）印度墨水，阴性染色，与常规光学显微镜结合使用，（ii）与细胞壁和胶囊的荧光染料共染色，然后进行共聚焦显微镜。最后,我们概述了如何手动测量胶囊直径，并提供了使用计算图像分析自动测量印度墨水染色样品直径的方案。
