Mozdziak P E, Fassel T A, Schultz E, Greaser M L, Cassens R G
Muscle Biology Laboratory, and Department of Anatomy, University of Wisconsin, Madison, 53706, USA.
Biotech Histochem. 1996 Mar;71(2):102-7. doi: 10.3109/10520299609117143.
A double fluorescence staining protocol was developed to facilitate computer based image analysis. Myofibers from experimentally treated (irradiated) and control growing turkey skeletal muscle were labeled with the anti-myosin antibody MF-20 and detected using fluorescein-5-isothiocyanate (FITC). Extracellular material was stained with concanavalin A (ConA)-Texas red. The cross-sectional area of the myofibers was determined by calculating the number of pixels (0.83 mu m(2)) overlying each myofiber after subtracting the ConA-Texas red image from the MF-20-FITC image for each region of interest. As expected, myofibers in the irradiated muscle were smaller (P < 0.05) than those in the non-irradiated muscle. This double fluorescence staining protocol combined with image analysis is accurate and less labor-intensive than classical procedures for determining the cross-sectional area of myofibers.
开发了一种双重荧光染色方案以促进基于计算机的图像分析。用抗肌球蛋白抗体MF-20对经实验处理（辐照）和对照生长的火鸡骨骼肌的肌纤维进行标记，并使用异硫氰酸荧光素（FITC）进行检测。细胞外物质用伴刀豆球蛋白A（ConA）-德克萨斯红染色。通过从每个感兴趣区域的MF-20-FITC图像中减去ConA-德克萨斯红图像后，计算覆盖每个肌纤维的像素数（0.83μm²）来确定肌纤维的横截面积。正如预期的那样，辐照肌肉中的肌纤维比未辐照肌肉中的肌纤维小（P<0.05）。这种双重荧光染色方案与图像分析相结合，对于确定肌纤维横截面积而言，比传统方法更准确且劳动强度更低。
