Baldwin W W, Bankston P W
Northwest Center for Medical Education, Indiana University School of Medicine, Gary 46408.
Appl Environ Microbiol. 1988 Jan;54(1):105-9. doi: 10.1128/aem.54.1.105-109.1988.
A new method is proposed to measure bacterial cells under growth conditions. Bacterial cells, suspended in their growth medium, were attached to a cover slip with poly-L-lysine. The cover slip was inverted and placed on a glass microscope slide. To prevent dehydration of the medium, the edges of the cover slip were sealed to the microscope slide with clear fingernail polish. The bacteria on the slide were then quickly photographed with a Leitz light microscope, using Nomarski optics. The photographic negatives were then projected at a standard distance through a lens system, and the projected images of the whole cells were outlined by hand onto graph paper. The profile images so transcribed onto the graph paper were in effect transverse sections of each of the cells. Using stereologic grid and point counting techniques, the area of the cell transverse section as well as the perimeter or circumference of the transverse section were estimated. Formulae were developed so that both the volume and surface area of the whole cell could be ascertained from these area and circumference measurements. Since the efficacy of any measurements of surface area and volume of microscopic rod-shaped bacterial cells could be questioned, macroscopic rod-shaped models were used to test the theory and formulae and to compare this method with other commonly used cell-sizing techniques. This technique could be used in any study of bacterial cell size or changes in cell size (e.g., osmotic shifts).
提出了一种在生长条件下测量细菌细胞的新方法。将悬浮在生长培养基中的细菌细胞用聚-L-赖氨酸附着在盖玻片上。将盖玻片倒置并放置在玻璃显微镜载玻片上。为防止培养基脱水，用透明指甲油将盖玻片边缘密封到显微镜载玻片上。然后使用诺马斯基光学系统，用徕卡光学显微镜对载玻片上的细菌快速拍照。然后将照相底片通过一个透镜系统在标准距离处投影，将整个细胞的投影图像手工勾勒在方格纸上。这样转录到方格纸上的轮廓图像实际上是每个细胞的横切面。使用体视学网格和点计数技术，估计细胞横切面的面积以及横切面的周长。开发了公式，以便根据这些面积和周长测量值确定整个细胞的体积和表面积。由于对显微镜下杆状细菌细胞表面积和体积的任何测量的有效性都可能受到质疑，因此使用宏观杆状模型来检验该理论和公式，并将该方法与其他常用的细胞大小测量技术进行比较。该技术可用于任何细菌细胞大小或细胞大小变化（如渗透变化）的研究。
