CN108276803A - 一种苏木素-伊红快速染色液及应用方法 - Google Patents
一种苏木素-伊红快速染色液及应用方法 Download PDFInfo
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Abstract
本发明公开了一种苏木素伊红快速染色液及其应用方法,属于生物技术领域。具体的说该苏木素伊红染色液包含改良苏木素染色液和新型伊红染色液。本发明提供的苏木素伊红染色液操作简便,染色均匀,染色后的细胞结构清晰,细胞着色鲜艳,易于临床病理诊断观察判断。
Description
技术领域
本发明涉及生物技术领域,特别涉及一种苏木素-伊红快速染色液,同时本发明还涉及该染色液应用方法。
背景技术
苏木素-伊红染色,简称HE染色,是病理组织切片最经常最广泛使用的一种常规染色法。苏木素容易被氧化,其氧化产物苏木红才是真正的燃料。习惯上称苏木素是碱性染料,实际上苏木红是一种酸性染料,只有苏木红和铝形成一种带正电荷的蓝色色精才是碱性的。带正电荷的蓝色色精和细胞核结合是通过正、负电荷的极性吸附来完成的,也就是说,带负电荷的脱氧核糖核酸根和带正电荷的蓝色色精进行极性吸附而完成染色。染色后必须进行分化和蓝化处理。所谓分化就是某些试剂,将过度染色的或不需要着色的组织成分上的颜色除去,所谓蓝化是指用明矾苏木素液深染细胞核后,通过盐酸乙醇分化,切片从酸性环境中转移至流水中使之变蓝的现象,本质上是染料苏木红加铝之间的结合或中断关系。一般来讲,在酸性环境中,使深蓝色的色精处于离子状态,此时为红色。这种现象称为色精形成中断,相反,红色离子状态的色精在碱性环境中处于结合状态,这种现象称色精形成,并呈现蓝色。
伊红Y是一种酸性红色胞浆性染料,含有1个醌型苯环的发色基和2个形成钠盐的酸性助色基,伊红Y溶于水中就能离解成带负电荷的色酸部分,即染料的有色部分;带正电荷的钠离子部分,即染料的无色部分。酸性染料组织成分呈弥漫性染色,这是由于组织成分的等电点一般都在3.6~5.2范围内,它们在弱酸性或弱碱性不可能进行极性吸附而完成染色,它们的染色可能是通过渗透作用或弥散作用而完成的,因此和组织成分结合是不牢固的,传统的苏木素-伊红染色过程中,易受组织固定、脱水、温度等因素的影响,导致以下问题:
1.细胞核着色较浅,轮廓不清并出现不同程度的片状发白区;
2.胞浆着色较淡,甚至不着色;
3.核浆共染,染色模糊,组织结构不清;
4.切片染色不均匀,出现点片状着色不良区;
5.切片不清晰,不透明;
6.切片染色不鲜艳,模糊。
发明内容
本发明为解决上述现有技术所存在的不足,提供一种快速染色的苏木素-伊红染色液及应用方法。该苏木素-伊红染色液配置简便,染色步骤简短,染后着色稳定,操作简单,染色效果显著。
具体来说:该苏木素-伊红染色液含有改良苏木素染色液和新型伊红染色液。
上述改良苏木素染色液,每升染色液其组分为:苏木色精0.5~5g,媒染剂2~20g,氧化剂0.01~1g,甲醇20~280ml,无水乙醇550~700ml,促染剂1~100ml,余量为纯化水。
上述新型伊红染色液,每升染色液其组分为:伊红Y醇溶5.5~8.2g,伊红Y水溶0.1~1.5g,橘黄G 0.1~2g,促染剂1~100ml,氨水15~200ml,无水乙醇750~950ml,余量为纯化水。
上述染色液中的媒染剂为十二水合硫酸铝钾、铁明矾中至少一种;所述氧化剂为黄色氧化汞、碘酸钠与高锰酸钾中至少一种;所述促染剂为氯化钠、硫酸钠、冰醋酸中至少一种。
本发明公开的新型伊红染色液使用水溶与醇溶伊红互补,水溶伊红易溶于水,溶液呈绿色荧光,使上皮细胞、肌肉纤维和细胞浆染色,醇溶性伊红能溶于碱,微溶于乙醇,吸附指示剂,不同的指示剂在不同的酸碱环境下呈现不同的颜色,使细胞浆、红细胞、肌肉、结缔组织、嗜伊红颗粒等被染成不同程度的红色或粉红色,与蓝色的细胞核形成鲜明对比,切片着色呈现明显的层次。
同时,本发明在传统的伊红染色液的成分基础上增加了橘黄G,能快速地作用于胞浆,缩短染色时间,对于宫颈、阴道上皮中非正常角化细胞和角化型鳞癌细胞的胞浆中都可以出现鲜艳的橘黄色。
染色液应用方法步骤为:
1)石蜡切片脱蜡2次,常温下每次1~3分钟;加热30~37℃,脱蜡1分钟;
2)无水乙醇1次,1~2分钟;
3)切片放置所述苏木素染液中1~2分钟;
4)取出玻片水洗30秒;
5)切片放置所述新型伊红染色液中5~10秒;
6)切片取出无水乙醇脱水1次,1分钟;
7)透明剂透明1~2分钟,取出切片封片,镜下阅片。
本发明公开的苏木素-伊红快速染色液的应用方法属于进行性染色,省去盐酸酒精分化过程,避免过度分化或者分化程度低影响判读结果。新型伊红染色液中添加碱性溶液氨水,使体系环境呈碱性,切片经过苏木素染色后可快速完成蓝化过程,同时胞浆着色。
附图说明
附图1本发明实施例一镜检图片。
附图2本发明实施例二镜检图片。
具体实施方式
下面用两个实施例对本发明涉及的苏木素-伊红快速染色液及应用方法做进一步说明。
实施例一:
改良苏木素染色液1,每升染色液其组分为:苏木色精2g,十二水合硫酸铝钾6g,碘酸钠0.06g,甲醇150ml,无水乙醇650ml,硫酸钠25ml,纯化水173ml。
新型伊红染色液1,每升染色液其组分为:伊红Y醇溶6.5g,伊红Y水溶0.8g,橘黄G0.6g,硫酸钠40ml,氨水90ml,无水乙醇800ml,纯化水68ml。
染色液染色步骤为:
1)石蜡切片脱蜡2次,常温下每次2分钟;
2)无水乙醇1次,2分钟;
3)切片放置所述苏木素染液中1分钟;
4)取出玻片水洗30秒;
5)切片放置所述新型伊红染色液中5秒;
6)切片取出无水乙醇脱水1次,1分钟;
7)透明剂透明2分钟,取出切片封片,镜下阅片。
实施结果:整个染色过程11分钟完成。完成后在显微镜下观察染色的效果:玻片细胞区域背景干净无杂质,细胞染色均匀,着色鲜艳,分色与染色效果明显,易于病理诊断判断。
实施例二:
改良苏木素染色液2,每升染色液其组分为:苏木色精1g,铁明矾2g,高锰酸钾0.03g,甲醇100ml,无水乙醇600ml,冰醋酸20ml,纯化水278ml。
新型伊红染色液2,每升染色液其组分为:伊红Y醇溶6.0g,伊红Y水溶0.03g,橘黄G0.3g,冰醋酸20ml,氨水40ml,无水乙醇800ml,纯化水136ml。
染色液染色步骤为:
1)石蜡切片脱蜡2次,加热37℃下每次1分钟;
2)无水乙醇1次,1分钟;
3)切片放置所述苏木素染液中1.5分钟;
4)取出玻片水洗30秒;
5)切片放置所述新型伊红染色液中5秒;
6)切片取出无水乙醇脱水1次,1分钟;
7)透明剂透明2分钟,取出切片封片,镜下阅片。
实施结果:整个染色过程9分钟完成。完成后在显微镜下观察染色的效果:玻片细胞区域背景干净无杂质,细胞染色均匀,着色鲜艳,分色与染色效果明显,易于病理诊断判断。
Claims (7)
1.一种苏木素-伊红快速染色液,其特征在于:含有改良苏木素液和新型伊红染色液。
2.根据权利要求1所述的苏木素-伊红快速染色液,其特征在于改良苏木素染色液,其每升染液其组分为:苏木色精0.5~5g,媒染剂2~20g,氧化剂0.01~1g,甲醇20~280ml,无水乙醇550~700ml,促染剂1~100ml,余量为纯化水。
3.根据权利要求1所述的苏木素-伊红快速染色液,其特征在于新型伊红染色液,每升染色液其组分为:伊红Y醇溶5.5~8.2g,伊红Y水溶0.1~1.5g,橘黄G 0.1~2g,促染剂1~100ml,氨水15~200ml,无水乙醇750~950ml,余量为纯化水。
4.根据权利要求2所述的改良苏木素染色液和权利要求3所述的新型伊红染色液,其特征在于组分中的促染剂为氯化钠、硫酸钠、冰醋酸中至少一种。
5.根据权利要求2所述的改良苏木素染色液,其特征在于组分中的媒染剂为十二水合硫酸铝钾、铁明矾中至少一种;所述氧化剂为黄色氧化汞、碘酸钠与高锰酸钾中至少一种。
6.根据权利要求3所述的新型伊红染色液,其特征在于染色液pH值为6.5~7.2。
7.根据权利要求1所述的苏木素-伊红快速染色液,其应用方法为:
1)石蜡切片脱蜡2次,常温下每次1~3分钟;加热30~37℃,脱蜡1分钟;
2)无水乙醇1次,1~2分钟;
3)切片放置所述苏木素染液中1~2分钟;
4)取出玻片水洗30秒;
5)切片放置所述新型伊红染色液中5~10秒;
6)切片取出无水乙醇脱水1次,1分钟;
7)透明剂透明1~2分钟,取出切片封片,镜下阅片。
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