Phelps M, Wilkins B S, Jones D B
Department of Pathology and Microbiology, Southampton General Hospital, UK.
Mol Pathol. 2000 Jun;53(3):159-61. doi: 10.1136/mp.53.3.159.
The isolation of p53 immunostain positive cells from histological sections for molecular genetic studies is a difficult task, especially if there are few positive cells. To eliminate contaminating DNA from p53 negative cells, which can obscure the results of molecular assays, a variation on the technique of immunohistoselective sequencing was developed. This is a highly selective approach, whereby immunostained sections of formalin fixed, paraffin wax embedded tissue are exposed to ultraviolet irradiation to damage the DNA in p53 negative cells. The DNA in positive cells remains unaffected because the dark immunostain protects their nuclei from ultraviolet light. Polymerase chain reaction single strand conformation polymorphism of samples enriched with p53 immunostain positive cells has shown that this method can produce pure samples of mutated DNA. The isolation of DNA from minority immunostain positive cells allows a wide range of molecular analyses to be carried out on these samples, which would otherwise be hampered by the problem of contaminating background cells.
从组织学切片中分离出p53免疫染色阳性细胞用于分子遗传学研究是一项艰巨的任务，尤其是当阳性细胞数量很少时。为了消除来自p53阴性细胞的污染DNA（其可能会掩盖分子检测的结果），人们开发了一种免疫组织选择性测序技术的变体。这是一种高度选择性的方法，通过将福尔马林固定、石蜡包埋组织的免疫染色切片暴露于紫外线照射下，以损伤p53阴性细胞中的DNA。阳性细胞中的DNA不受影响，因为深色的免疫染色可保护其细胞核免受紫外线照射。对富含p53免疫染色阳性细胞的样本进行聚合酶链反应单链构象多态性分析表明，该方法可以产生纯的突变DNA样本。从小部分免疫染色阳性细胞中分离DNA，使得对这些样本能够进行广泛的分子分析，否则这些分析会因背景污染细胞的问题而受到阻碍。
