Boland C R, Bresalier R S, Roberts J A
GI Research Laboratory, Veterans Administration Medical Center, San Francisco, California.
J Histochem Cytochem. 1988 Oct;36(10):1329-34. doi: 10.1177/36.10.3138308.
We have developed a technique for quantitation of binding of fluorescent lectins to glycoconjugates in specimens of tumors derived from cultured human colorectal cancer cells. Tumor cells were injected subcutaneously into nude mice, giving rise to xenografts that resemble primary human colorectal cancers. The tumors were extracted with saline and were subjected to dialysis and lyophilization. Standardized amounts of the tumor extract were then incubated with fluorescent lectins and subjected to gel permeation liquid chromatography to separate lectin bound to high molecular weight glycoproteins from free (unbound) lectin, and were quantitated using a spectrofluorometer. This assay permitted quantitative measurement of the lectin bound to high molecular weight glycoconjugates such as mucin. The results of this assay were compared with the standard histochemical assessment of tissue labeling by fluorescent lectins. A close correlation between the two techniques was found, especially when little or no labeling was present. Greater variations were observed at higher levels of labeling. The quantitative assay confirms that lectins bind to high molecular weight mucin-type glycoconjugates on fixed sections of tumors, and supports the use of semi-quantitative histochemical assessments of tissue labeling.
我们已经开发出一种技术，用于定量检测荧光凝集素与源自培养的人结肠癌细胞的肿瘤标本中的糖缀合物的结合。将肿瘤细胞皮下注射到裸鼠体内，形成类似于原发性人结肠癌的异种移植瘤。用生理盐水提取肿瘤，然后进行透析和冻干。然后将标准化量的肿瘤提取物与荧光凝集素一起孵育，并进行凝胶渗透液相色谱，以将与高分子量糖蛋白结合的凝集素与游离（未结合）凝集素分离，并用荧光分光光度计进行定量。该测定法允许定量测量与高分子量糖缀合物（如粘蛋白）结合的凝集素。将该测定结果与荧光凝集素对组织标记的标准组织化学评估进行比较。发现这两种技术之间密切相关，尤其是在几乎没有标记或没有标记的情况下。在较高标记水平下观察到更大的差异。定量测定证实凝集素与肿瘤固定切片上的高分子量粘蛋白型糖缀合物结合，并支持对组织标记进行半定量组织化学评估。
