利用不同染料复合染色判定家蚕血细胞的存活状况

家蚕有5种血细胞,可以通过吖啶橙(acridine orange,AO)和碘化丙啶(propidium iodide,PI)染色来区分。溴化噻唑蓝四氮唑3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide,MTT]是一种广泛应用于细胞增殖和活细胞鉴定的染色剂。通过对家蚕血细胞进行复染,即先用MTT染色,再用AO和PI染色,发现以下2种情况:很多颗粒细胞、浆细胞和所有的原血球细胞不能被MTT染色,但AO染色显示为活的细胞;被MTT染色的细胞表面有紫色纤维状物伸出,除了一部分具有伪足的颗粒细胞外,所有被MTT染色的血细胞也被PI染色,细胞核是红色。因此,利用这3种染料对家蚕血细胞复合染色来判断细胞的存活与死亡,需要仔细分析并分别对待。

Using Different Staining Agents to Identify the Living Status of Circulating Hemocytes in the Silkworm, Bombyx mori

Bombyx mori has 5 types of circulating hemocytes that can be clearly identified after being stained by acridine orange(AO) and propidium iodide(PI).3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide(MTT) is another type of staining agent that has been extensively used to study cell proliferation and to identify living cells.Bombyx mori hemocytes undergone multiple staining,i.e.staining with MTT first and then with AO and PI,showed two patterns.One was that a majority of granulocytes and plasmatocytes and all prohemocytes could not be stained by MTT but were shown to be living cells from AO staining.The other was that the MTT stained hemocytes had a lot of purple fiber-like structure extending from cell surface.All MTT-stained hemocytes but some granulocytes with pseudopods could also be stained by PI according to their nuclei with red fluorescence.Therefore,careful analysis and discriminative treatment should be conducted when multiple staining with these three staining agents are used to tell whether a cell is living or dead.