本文關(guān)鍵詞：免疫組織化學(xué)染色防脫片幾種方法的比較及PCNA、MMP-7、VEGF、Cath-D和HSP70在食管癌中的表達(dá)及臨床意義　出處：《河北醫(yī)科大學(xué)》2005年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 粘附劑 免疫組織化學(xué)染色 防脫片 APES 多聚賴氨酸 白乳膠 食管癌 細(xì)胞增殖期核蛋白 基質(zhì)金屬蛋白酶-7 血管內(nèi)皮生長因子 組織蛋白酶D 熱休克蛋白70 預(yù)后

【摘要】：目的: 在今天的免疫學(xué)實驗研究及病理診斷中,免疫組織化學(xué)(免疫組化)是一種應(yīng)用較廣的標(biāo)記免疫技術(shù)。免疫組化具有操作簡便,結(jié)果易于觀察,特異性較高等優(yōu)點,但免疫組化染色結(jié)果并不十分不穩(wěn)定,影響因素較多。研究發(fā)現(xiàn),玻片的清潔程度,不同的抗原修復(fù)方法(微波、高壓鍋、水浴煮沸和酶消化),緩沖液的pH 值,是否經(jīng)高壓處理切片以及不同公司的試劑類型對免疫組化染色都有影響。有些學(xué)者致力于探討免疫組化染色的影響因素,制定出統(tǒng)一的操作規(guī)范和評定標(biāo)準(zhǔn),以保證免疫組化的正確使用。石蠟切片在免疫組化染色中的脫片問題由來已久。切片經(jīng)抗原修復(fù)和緩沖液的長時間浸泡后,往往會部分或全部從載玻片上脫落,影響染色結(jié)果的判斷。不同的粘附劑和粘附劑不同的涂膠方法防脫片效果不盡相同。我們在工作中也經(jīng)常遇到石蠟切片脫落問題,為此本實驗對三種粘附劑即APES(氨醛三已氧基硅烷),多聚賴氨酸和白乳膠的染色結(jié)果和防脫片效果作了比較;又進(jìn)一步對多聚賴氨酸的涂膠方法作了比較和改進(jìn),成功地總結(jié)出一種重復(fù)性好的多聚賴氨酸涂膠法。 方法: 本實驗用3 種粘附劑APES、多聚賴氨酸、白乳膠涂膠,APES 涂膠按說明書,白乳膠涂膠參照文獻(xiàn)。多聚賴氨酸涂膠除了按說明書用浸泡法以外,還對其涂膠條件做 了探討,0.01%多聚賴氨酸不同體積(40μL, 80μL, 120μL,160μL)單次涂膠;同量多聚賴氨酸不同濃度單次涂膠(0.02%多聚賴氨酸40μL、0.0133%多聚賴氨酸60μL、0.01%多聚賴氨酸80μL);0.01%多聚賴氨酸80μL 單次涂膠和雙次涂膠;0.01%多聚賴氨酸80μL 單次涂膠后37℃保存的影響(0.01%多聚賴氨酸80μL 單次涂膠后當(dāng)日貼片或在37℃放置4 周后再貼片)。檸檬酸加熱做抗原修復(fù)以后進(jìn)行免疫組化染色,HPIAS-1000 高清晰度彩色病理系統(tǒng)測量切片染色強(qiáng)度;肉眼觀察脫片情況。 結(jié)果: 1 不同粘附劑染色強(qiáng)度和防脫片效果的比較特異性顯色OD 均值,APES 組為0.1980±0.0120,多聚賴氨酸浸泡法組為0.2020±0.0082,白乳膠組為0.2580±0.0164。非特異性顯色OD 均值,APES 組為0.0105±0.0010,多聚賴氨酸浸泡法組為0.0106±0.0009,白乳膠組為0.0298±0.0027。APES 組和多聚賴氨酸浸泡法組特異性顯色、非特異性顯色的差別均沒有顯著性(P0.05);白乳膠組特異性顯色、非特異性顯色均顯著強(qiáng)于APES 組(P0.01)和多聚賴氨酸浸泡法組(P0.01)。特異性顯色OD 均值除以非特異性顯色OD 均值,APES 組為18.81,多聚賴氨酸浸泡法組為19.08,白乳膠組為8.66。三種粘附劑的完整貼片百分率分別為APES 組55%,多聚賴氨酸浸泡法組57.5%,白乳膠組50%,三組均不能獲得高比例的完整貼片。 2 多聚賴氨酸不同涂膠方法防脫片效果的比較 1)相同濃度不同體積多聚賴氨酸的比較0.01%多聚賴氨酸40μL, 80μL, 120μL, 160μL 四種不同體積單次涂膠的完整貼
[Abstract]:Objective: in today's study on immunological function and pathological diagnosis, immunohistochemical (IHC) staining technique is a widely used. Immunohistochemistry has the advantages of simple operation, easy observation results, the specificity is relatively high, but the results of immunohistochemical staining and not very unstable factors research found that the clean degree of slide, different methods of antigen retrieval (microwave, pressure cooker, water boiling and enzyme digestion), buffer pH value, whether by high pressure treatment section and different company type reagents on immunohistochemical staining are affected. Some scholars committed to the study of immunohistochemical staining effect factors, formulate uniform operating standards and evaluation standards, to ensure the correct use of immunohistochemistry in paraffin. Immunohistochemical staining in the unfalling sections of the long-standing problem. The antigen retrieval and buffer time Between after soaking, often partly or wholly from the slide off, affect the judgement of the staining results. Gluing method of adhesion agent and adhesion agent of different anti stripping effect is not the same. In our work we often encounter paraffin shedding problem, this experiment of three kinds of adhesive APES (ammonia three aldehyde silane), poly lysine staining and white latex and anti off effect were compared; and further to the polylysine coating method of acid ammonia were compared and improved, successfully concluded a reproducible polylysine coating method.
Methods: 3 adhesives APES, polylysine and white latex were applied to the experiment, and APES coating was applied according to the instructions. The white latex was coated with reference literature.
On 0.01% poly-L-lysine of different volume (40 L, 80 L, 120 L, 160 L) gelatinizedonce; the same amount of poly-L-lysine of different concentrations of gelatinizedonce (0.02% poly lysine 40 L 0.0133% poly lysine 60 L 0.01% poly lysine 80 L); 0.01% poly lysine 80 L single coating and double coating; 0.01% poly lysine save 37 C 80 L gelatinizedonce after 0.01% (poly lysine 80 L gelatinizedonce after the patch or in 37 C placed 4 weeks before the patch). After antigen repairing withcitric acid by immunohistochemical staining, HPIAS-1000 high-resolution color measurement system pathological staining intensity; the unfalling observed with naked eye.
Result:
1 different adhesive strength and anti staining effect and comparison of specific color OD mean, APES group is 0.1980 + 0.0120, poly-L-lysine dipping group is 0.2020 + 0.0082, white latex color group mean OD was 0.2580 + 0.0164. nonspecific, APES group is 0.0105 + 0.0010, poly lysine ammonia acid immersion group is 0.0106 + 0.0009, white latex group is 0.0298 + 0.0027.APES group and poly-L-lysine dipping group specific color, non-specific color differences were not significant (P0.05); white latex specific color, non-specific staining was significantly stronger than that of APES group (P0.01) and poly-L-lysine dipping group (P0.01). The color OD value divided by the non-specific staining of OD mean specificity, APES group was 18.81, poly-L-lysine dipping group was 19.08, white latex group complete patch percentage of 8.66. of three kinds of adhesion agent were 55% in group APES, poly-L-lysine dipping Group 57.5%, white latex group 50%, three groups were not complete patch to obtain a high proportion.
Comparison of the effect of more than 2 polylysine with different coating methods
1) the comparison of the same concentration of polylysine with different volumes of polylysine 0.01% polylysine 40 mu L, 80 mu L, 120 micron L, 160 L four different volume single coating adhesive

【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2005
【分類號】：R735.1;R392

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