WO2013120394A1 - 一种检测或诊断***癌的试剂盒 - Google Patents

一种检测或诊断***癌的试剂盒 Download PDF

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WO2013120394A1
WO2013120394A1 PCT/CN2013/000118 CN2013000118W WO2013120394A1 WO 2013120394 A1 WO2013120394 A1 WO 2013120394A1 CN 2013000118 W CN2013000118 W CN 2013000118W WO 2013120394 A1 WO2013120394 A1 WO 2013120394A1
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catenin
antibody
prostate cancer
kit
buffer
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PCT/CN2013/000118
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French (fr)
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曾燕
钱泽
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昂科生物医学技术(苏州)有限公司
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
    • G01N33/57407—Specifically defined cancers
    • G01N33/57434—Specifically defined cancers of prostate
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing

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  • the present invention relates to a kit, and more particularly to an agent for detecting or diagnosing prostate cancer.
  • Prostate cancer is one of the common cancers and ranks second among male lethal cancers (Jemal, A., Siegel R, Xu J, Ward E. Cancer statistics, 2010. CA Cancer J Clin 2010. 60 ( 5): 277-300; Lu Q, Zhang J and Chen YH. Prostate cancer cell growth and death: complex roles of pro- and anti-oncogenic protein signaling. 2009. In: Handbook of Prostate Cancer Cell Research (Editor: Alan T Meridith). Nova Science Publishers, Inc. 431-447; Hessels D and Schalken JA. The use of PCA3 in the diagnosis of prostate cancer. 2009. Nature Reviews Urology. 6: 255-261). Due to the complexity of prostate cancer performance, early diagnosis of prostate cancer is difficult.
  • PSA serum prostate specific antigen
  • DRE digital rectal examination
  • TRUS transrectal ultrasound
  • Urine biomarker detection is very attractive because it provides a simple, Non-invasive Early prostate cancer screening methods allow a wide range of people to be screened. In recent years, great progress has been made in the discovery of urine markers for prostate cancer. A urine biomarker was validated using PCA3 (prostate cancer gene 3) non-coding m NA and TMPR S2:ERG fusion gene technology. However, they need to take urine after prostate massage or the positive value is lower.
  • PCA3 prostate cancer gene 3
  • ⁇ -Catenin/NPRAP/Neurojungin ( ⁇ -cyclonectin/NPRAP/neuronectin) is an adhesion junction protein in the ⁇ -catenin superfamily (Lu, Q, Paredes M, Medina M, Zhou J, Cavallo R, Jifer Biol, 1999. 144(3): 519-532), originally identified as a specific protein in brain tissue.
  • ⁇ -catenin mRNA is overexpressed in prostate cancer compared with benign prostatic hyperplasia (Lu Q, Dobbs LJ, Christopher WG, Lanford GW, Revelo MP, Shappell S and Chen YH.
  • NPRAP Plakophilin-related armadillo protein
  • tissue microarray TMA to analyze 90 specimens of prostate cancer after prostatectomy and 90 specimens of normal prostate tissue, 92% of patients with Cap showed strong (46/72) or moderate (20/72) ⁇ - Catenin staining (immune score ⁇ 2, only 6 of the 72 specimens (8%) were negative for the immune score or ⁇ 2. 65 of the 65 benign specimens (75%) had an immune score ⁇ 2, and 65 of the benign specimens were Six patients (9%) had an immune score equal to 2, and 10 of 15 65 (15%) benign specimens had an immune score higher than 2. These data indicate that the clinical sensitivity of ⁇ -catenin alone is 91.7%.
  • a kit for diagnosing and diagnosing prostate cancer which mainly comprises ⁇ -catenin adsorbing antibody, coating buffer, washing buffer, blocking buffer, ⁇ -catenin detecting antibody, and enzyme labeling antibody.
  • the invention also provides a simple and easy method A kit for detecting or diagnosing a prostate, which mainly comprises a ⁇ -catenin adsorbing antibody, a coating buffer, a washing buffer, a blocking buffer, a ⁇ -catenin detecting antibody, and a nano quantum dot label.
  • the standard is preferably a human ⁇ -catenin standard protein.
  • ⁇ -catenin adsorption resistance The body may be rabbit anti-human, mouse anti-human, chicken anti-human or other ⁇ -catenin antibody, and the ⁇ -catenin antibody is diluted to a protein content of l ⁇ 10 ( ⁇ g / ml) when used.
  • the coating buffer may be For carbonate or dPBS or other buffer.
  • the wash buffer is preferably detergent-containing dPBS or other buffer.
  • the ⁇ -catenin detecting antibody may be rabbit anti-human, mouse anti-human, chicken anti-human or other ⁇ -catenin antibody.
  • the ⁇ -catenin recognition antibody recognizes a different ⁇ -catenin protein sequence from the ⁇ -catenin detection antibody.
  • the invention further relates to the use of a ⁇ -catenin protein for the preparation of a preparation for detecting or diagnosing prostate cancer.
  • the kit provided by the invention can detect ⁇ -catenin protein which is highly correlated with prostate cancer by ELISA in the urine of the patient, and the ELISA method is mature, sensitive and specific, and non-invasive to the patient, Can be repeated, the patient is willing to accept. Further, the nanoquantum dot (QD) labeling method has a small sample size, and the antibody used in the ELISA method is combined with QD to more sensitively detect changes in the ⁇ -catenin protein in the urine for diagnosis of prostate cancer.
  • QD nanoquantum dot
  • Figures 1A-1B show the ⁇ -catenin protein in urine by ELISA.
  • Figure 1A shows the purified standard protein ⁇ -catenin (ng/ml)
  • Figure 1B shows the ⁇ -catenin enzyme reaction in urine.
  • Figure 2 shows the detection of ⁇ -catenin protein in urine by nanometer quantum dot labeling.
  • the present invention provides a simple and easy kit for detecting or diagnosing prostate cancer, which mainly comprises ⁇ -catenin adsorption antibody, coating buffer, washing buffer, blocking buffer, ⁇ -catenin detection antibody, enzyme Target antibody. Or mainly comprising ⁇ -catenin adsorption antibody, coating buffer, washing buffer, blocking buffer, ⁇ -catenin detection antibody, nano quantum dot labeling.
  • the kit also contains standards and a negative control.
  • the following detailed description uses the kit of the present invention to detect or diagnose prostate cancer.
  • the human ⁇ -catenin standard protein of the present invention is a purified recombinant human ⁇ -catenin protein, and an anti-human ⁇ -catenin antibody has been disclosed in the following literature: Paffenholz R, Franke WW. Identification and localization of a neurally expressed member of the Plakiglobin/armadillo multigene family. Differentiation. 1997 Aug;61(5):293-304 ; Lu Q, Paredes M, Medina M, Zhou J, Cavallo R, Peifer M Orecchio L, Kosik KS. delta-catenin, an adhesive junction -associated protein which promotes cell scattering. J Cell Biol. 1999 Feb 8; 144(3): 519-32.
  • ⁇ -catenin antibody Dilute the ⁇ -catenin antibody to a protein content of 1 to 10 ( ⁇ g / ml) with carbonate or dPBS coating buffer. Add 100 ⁇ l to the reaction well of each polystyrene plate, overnight at 4 ° C. On the next day, the solution in the well was discarded, washed 5 times with washing buffer, and washed by a plate washer.
  • Blocking buffer (Blocking buffer)
  • Step 3 Load the sample
  • Negative control Normal healthy adult male urine specimens.
  • Prostate cancer urine specimens - 37 patients with prostate cancer aged 50 to 80 were diagnosed by urology ultrasound and pathologically diagnosed Gleason grade 7 ⁇ 10. Their urine is stored at -20 degrees after collection.
  • Each specimen was given 100 ⁇ l of urine and incubated at 37 ° C for 2 hours. Wash the plate.
  • Urine specimens can also be centrifuged at low temperature and high speed (14,000) for 15 minutes, and the sediment is filled with 3 wells and the urine is cleared to three wells.
  • Step 4 Add ⁇ -catenin detection antibody diluted with Block solution, 50 ⁇ l per well. Wash the plate.
  • Step 5 Add the enzyme-labeled antibody (goat anti- or other anti-HRP) (Sigma, USA). Add freshly diluted anti-HRP enzyme-labeled antibody (Sigma, USA) to each well. After incubating for 1 hour at room temperature, the plate washer was washed.
  • TMB (Sigma T0440) was added to each well and stored at room temperature in the dark. Or Shanghai Kehua Company coloring liquid A and B liquid.
  • the reaction was terminated by adding 3 ⁇ 4 S0 4 per well.
  • Step 8 Result determination
  • the reader reads the board.
  • Example 2 Determination of ⁇ -catenin protein in urine by nanometer quantum dot labeling
  • Steps 1 to 4 are the same as in the first embodiment.
  • Step 5 Nano-quantum dot labeling is combined with the anti-human ⁇ -catenin antibody of step 4 using Evitas Technologies (Troy, New York, USA) Evitags nano-quantum dot water-soluble.
  • EDC (1 -ethyl-3-[3-dimethylaminopropyl] carbodiimide) and sulfo-NHS were used to form an activity on the surface of nano quantum dots. After the excess EDC and sulfo-NHS were removed by a filter, an anti-human ⁇ -catenin antibody was added to form a covalent bond with the nano quantum dots.
  • Step 6 Result determination
  • nano-quantum dot labeling experiments show that nano-quantum dots (QD) combined with ⁇ -catenin antibodies can also measure ⁇ -catenin protein in solution.
  • QD nano-quantum dots
  • ⁇ -catenin antibody binding enzymes or binding to nano quantum dots can detect changes in ⁇ -catenin protein which is highly correlated with prostate cancer, and thus can be used for diagnosis of prostate cancer.
  • ⁇ -catenin antibodies bind to or bind to nano quantum dots as long as ⁇ -catenin proteins are recognized by ⁇ -catenin antibodies or ⁇ -catenin antibodies and nano quantum dot conjugates. Can be used for testing.
  • the scope of protection of the present invention is defined by the scope of the claims, and the scope of the invention is defined by the appended claims.

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Abstract

一种检测或诊断***癌的试剂盒，其包含δ-catenin吸附抗体、包被缓冲液、洗涤缓冲液、封闭缓冲液、δ-catenin检测抗体和酶标抗体，或包含δ-catenin吸附抗体、包被缓冲液、洗涤缓冲液、封闭缓冲液、δ-catenin检测抗体和纳米量子点标记。该试剂盒能够检测出尿液中δ-catenin蛋白的变化以用于***癌诊断，具有非侵袭性，高敏感性和特异性的优点。

Description

一种检测或诊断***癌的试剂盒 技术领域 本发明涉及一种试剂盒， 具体涉及一种检测或诊断***癌的试剂 说
品.。 书
背景技术 ***癌（PCa)是常见癌症之一，在男性致死癌症中排位第二 （Jemal, A., Siegel R, Xu J, Ward E. Cancer statistics, 2010. CA Cancer J Clin 2010. 60 (5): 277-300； Lu Q, Zhang J and Chen YH. Prostate cancer cell growth and death: complex roles of pro- and anti-oncogenic protein signaling. 2009. In: Handbook of Prostate Cancer Cell Research (Editor: Alan T. Meridith). Nova Science Publishers, Inc. 431-447; Hessels D and Schalken JA. The use of PCA3 in the diagnosis of prostate cancer. 2009. Nature Reviews Urology. 6: 255-261)。 由于***癌表现的复杂性， 早期诊断***癌是困难的。 目 前临床广泛采用的筛选方法是检测血清***特异性抗原 （PSA) , 数字 直肠检查 （DRE) 和经肠的超声检测 （TRUS)。 但 PSA 的关键统计实验 表明 PSA对 PCa的阳性预测值仅 34%， 在灰区 4-l(^g/L的患者有 25%有 隐匿的 PCa。 PSA浓度 < 4 g/L的男性有 15%有 PCa。 而良性***增生 (BPH) , 其 PSA也可以升高， 可以导致不必要的病理活组织检查和其它 昂贵且不舒服的侵袭性检查介入。作为现在临床采用的唯一个***癌血 清标志物， PSA有明显的不足之处 (Hessels D and Schalken JA. The use of PCA3 in the diagnosis of prostate cancer. 2009. Nature Reviews Urology. 6: 255-261)。 因此寻找一个简单易行的， 使 PCa特异性、 敏感性、 检出率大 大增加的标志物， 将对 PCa的早期发现、 诊断和治疗产生深远的影响。尿 液生物标记检测是非常有吸引力的， 因为这可提供一个简单， 非侵袭性的 早期***癌检查方法， 可以使广泛的人群接受筛选。 近年来， 有关前列 腺癌尿液标记物的发现有了很大的进展。 用 PCA3 (***癌基因 3 ) 非 编码 m NA和 TMPR S2:ERG 融合基因技术验证了一个尿液生物标记。 可是它们需要***按摩后取尿否则阳性值较低。其它一些尿液生物标记 目前正在研究中， 包括 PCADM-1 , 肌氨酸， Engrailed基因， 微染色体 5 蛋白， ***特异膜抗原等。一个理想的尿液生物标记必须满足一些重要 的标准， 即除了能够区别正常组织和癌组织， 对***癌病理有科学的支 持， 并且其检测方法应该简单无痛， 应该使临床医生容易对其进行解释。
δ-Catenin/NPRAP/Neurojungin ( δ-环连蛋白/ NPRAP/神经连接蛋白） 是 β-catenin超家族中的一个粘附连接蛋白 （Lu, Q, Paredes M, Medina M, Zhou J, Cavallo R, Peifer M, Orecchio L, Kosik KS. delta-catenin, an adhesive junction-associated protein which promotes cell scattering. J Cell Biol, 1999. 144(3): 519-532), 最初鉴定为脑组织中的特异蛋白。 然而研究表明， 与良 性***增生相比， ***癌的 δ-catenin mRNA过度表达 （Lu Q, Dobbs LJ, Christopher WG, Lanford GW, Revelo MP, Shappell S and Chen YH. Increased expression of δ-catenin/neural plakophilin-related armadillo protein (NPRAP) is associated with the downregulation and redistribution of E-cadherin and l20ctn in human prostate cancer. 2005. Human Pathology. 36: 1037-1048； Burger, MJ Tebay MA, Keith PA, Samaratunga HM, Clements J, Lavin MF et al" Expression analysis of delta-catenin and prostate-specific membrane antigen: their potential as diagnostic markers for prostate cancer. Int J Cancer. 2002. 100(2): 228-237)。 组织微阵列研究表明， ***上皮内瘤 变中 (PIN)S-catenin表达开始升高，而且与 Gleason评分增高相一致，表明 δ-catenin是***癌进展的一个潜在的指标 （Lu Q, Dobbs LJ, Christopher WG, Lanford GW, Revelo MP, Shappell S and Chen YH. Increased expression of δ-catenin/neural plakophilin-related armadillo protein (NPRAP) is associated with the downregulation and redistribution of E-cadherin and l20ctn in human prostate cancer. 2005. Human Pathology. 36: 1037-1048； Lu Q and Chen YH. Method of detecting cancer using delta-catenin. United States Patent: 7,445,906. 2008)。
用组织微列阵 TMA分析 90例***切除后的***癌组织标本和 90例正常***组织标本， 92%的 Cap病人标本显示出强的 (46/72 ) 或中 度 （20/72) δ-catenin染色（免疫评分≥2， 72例标本中只有 6例（8%)免疫 评分阴性或 <2。 65例良性标本中有 49例 (75%)免疫评分<2， 65例良性 标本中有有 6例 （9%) 免疫评分等于 2， 65例良性标本中有 10例 (15%) 免疫评分高于 2。 这些资料表明临床上如果单独用 δ-catenin作为评价， 其 敏感性为 91.7% , 特异性为 75.4% (Lu Q, Dobbs LJ, Christopher WG, Lanford GW, Revelo MP, Shappell S and Chen YH. Increased expression of δ-catenin/neural plakophilin-related armadillo protein (NPRAP) is associated with the downregulation and redistribution of E-cadherin and l20ctn in human prostate cancer. 2005. Human Pathology. 36: 1037-1048)。 最近的一个关于人 尿液沉淀物的研究显示在人的无细胞尿液***小体中可检测到 δ-catenin ， PCa病人尿中 δ-Catenin显著增加 (P < 0.0005)(8)。 这些研究提 出一个为 PCa病人非侵袭检测的可能性 （Lu Q, Zhang J, Allison R, Gay H, Yang WX, Bhowmick N, Frelix G, Shappell S, Chen YH. Identification of extracellular -catenin accumulation for prostate cancer detection. 2009. The Prostate. 69(4) :411-418) 。 发明内容 本发明的目的在于提供一种简单易行的检测或诊断诊断***癌的 试剂盒， 其主要包含 δ-catenin吸附抗体、 包被缓冲液、 洗涤缓冲液、封闭 缓冲液、 δ-catenin检测抗体、 酶标抗体。 本发明还提供一种简单易行的检测或诊断诊断***的试剂盒， 其主 要包含 δ-catenin吸附抗体、包被缓冲液、洗涤缓冲液、封闭缓冲液、 δ-catenin 检测抗体、 纳米量子点标记。
在本发明提供的上述试剂盒中还包含标准品和阴性对照。
其中所述标准品优选为人 δ-catenin标准蛋白。 所述 δ-catenin吸附抗 体可为兔抗人，鼠抗人，鸡抗人或其它 δ-catenin抗体，使用时所述 δ-catenin 抗体稀释至蛋白质含量为 l〜10(^g / ml。 所述包被缓冲液可为碳酸盐或 dPBS或其它缓冲剂。
所述洗涤缓冲液优选为含洗涤剂的 dPBS或其它缓冲剂。
其中所述 δ-catenin检测抗体可为兔抗人， 鼠抗人， 鸡抗人或其它 δ-catenin抗体。 优选 δ-catenin识别抗体与 δ-catenin检测抗体识别不同 δ-catenin蛋白序列。
本发明还涉及 δ-catenin蛋白在制备检测或诊断***癌的制剂中的 应用。
本发明提供的试剂盒， 能够从病人尿液中用 ELISA方法检测与前列 腺癌成高度相关性的 δ-catenin蛋白， 而且 ELISA方法成熟， 敏感性和特 异性较高， 对病人是非侵袭性的， 能够重复采取， 病人乐于接受。 进一歩 地， 纳米量子点 （QD) 标记法取样量小， 将 ELISA方法中所用的抗体与 QD结合， 能够更灵敏地检测尿液中 δ-catenin蛋白的变化， 从而用于诊断 ***癌。
为让本发明的上述和其它目的、 特征和优点能更明显易懂， 下文特举 较佳实施例， 并配合附图， 作详细说明如下。 附图说明 图 1A-1B是 ELISA法检测尿液中的 δ-catenin蛋白， 其中图 1A表示纯化 标准蛋白 δ-catenin (ng/ml) ， 图 1B是尿液 δ-catenin酶联反应。
图 2是纳米量子点标记法检测尿液中 δ-catenin蛋白。 具体实施方式 本发明提供一种简单易行的检测或诊断***癌的试剂盒，其主要包 含 δ-catenin吸附抗体、 包被缓冲液、 洗涤缓冲液、 封闭缓冲液、 δ-catenin 检测抗体、 酶标抗体。 或者主要包含 δ-catenin吸附抗体、 包被缓冲液、 洗 涤缓冲液、 封闭缓冲液、 δ-catenin检测抗体、 纳米量子点标记。 所述试剂盒还包含标准品和阴性对照。
下述详细说明采用本发明的试剂盒检测或诊断***癌。
本发明所述的人 δ-catenin标准蛋白是纯化的重组人 δ -catenin蛋白、 抗人 δ-catenin 抗体已公开于下述文献： Paffenholz R, Franke WW. Identification and localization of a neurally expressed member of the plakoglobin/armadillo multigene family. Differentiation. 1997 Aug;61(5):293-304; Lu Q, Paredes M, Medina M, Zhou J, Cavallo R, Peifer M Orecchio L, Kosik KS. delta-catenin, an adhesive junction-associated protein which promotes cell scattering. J Cell Biol. 1999 Feb 8; 144(3):519-32.
实施例 h ELISA法检测尿液中 δ-catenin蛋白
步骤 1: 包被 (Coat plate)
用碳酸盐或 dPBS包被缓冲液将 δ-catenin吸附抗体稀释至蛋白质含量 为 l〜10(^g / ml。 在每个聚苯乙烯板的反应孔中加 100微升， 4°C过夜。 次 日， 弃去孔内溶液， 用洗涤缓冲液洗 5次， 洗板机洗涤。
1) 包被缓冲液 （碳酸盐缓冲液） ：
NaC03 1.59克
NaHC03 2.93克
加蒸馏水至 1000ml
或 Dulbecco's PBS (Invitrogen)
2) 洗涤缓冲液：
KH2P04 0.2克
Na2HP04 12H20 2.9克
NaCl 8.0克
KC1 0.2克
Tween-20 0.05 % 0.5ml
加蒸馏水至 1000ml 步骤 2: 封闭
每孔加封闭缓冲液 200微升， 室温下孵育 1小时后。 用洗板机洗涤。 封闭液： （Blocking buffer)
牛血清白蛋白 （BSA) 2克
加洗涤缓冲液至 100ml。
步骤 3: 加样
标准品： 人 δ-catenin标准蛋白。
阴性对照： 正常健康成年男性尿液标本。
***癌症尿液标本—— 37例年龄 50〜80的***癌病人经泌尿科超 声初诊和病理确诊 Gleason分级 7〜10。 他们的尿液收集后在 -20度保存。
每个标本尿液各加 100微升， 37°C温育 2小时后。 洗板。
尿液标本也可低温高速（14,000)转离心 15分钟， 加样沉淀物 3孔， 尿 上清三孔。
步骤 4: 加 δ-catenin检测抗体用 Block液稀释， 每孔 50微升。 洗板。 步骤 5: 加酶标抗体 (羊抗或其它抗 -HRP) (美国 Sigma公司） 于各反应孔中，加入新鲜稀释的抗 -HRP酶标抗体（美国 Sigma公司）。 室温孵育 1小时后， 洗板机洗涤。
步骤 6: 显色
于各反应孔中加 TMB ( Sigma T0440) , 室温、 避光保存。 或上海科 华公司显色液 A和 B液。
步骤 7: 终止反应
每孔加 ¾S04,终止反应。
步骤 8: 结果判定
读板机读板。
实验表明， 尿液中可用双抗体夹心酶联免疫法测到至少 2ng 以下 S-catenin。 如图 1A-1B所示， 其中其中图 1A表示纯化标准蛋白 δ-catenin (ng/ml) ， 图 1B是尿液 δ-catenin酶联反应。 从图 1B中可以看到， 前列 腺癌病人尿液 δ-catenin可达正常人的 3倍。 运用 48例***癌病人尿液 与正常人试验鉴定敏感性可达 65〜85%， 特异性可达 90%以上。
实施例 2: 纳米量子点标记法检测尿液中 δ-catenin蛋白
步骤 1至步骤 4同实施例 1。
步骤 5: 纳米量子点标记与步骤 4的抗人 δ-catenin抗体结合 采用 Evident Technologies (Troy, 美国纽约）的 Evitags 纳米量子点水 溶 。 用 EDC (1 -ethyl-3-[3-dimethylaminopropyl] carbodiimide)和 sulfo-NHS 在纳米量子点表面形成活性。 在多余的 EDC and sulfo-NHS 用过滤器去除后， 加入抗人 δ-catenin抗体与纳米量子点形成共价结合。
步骤 6: 结果判定
如图 2 所示， 纳米量子点标记实验显示， 纳米量子点 （QD) 结合 δ-catenin抗体也能测定溶液中的 δ-catenin蛋白。
以上的这些试验都证明用 δ-catenin抗体结合酶联或结合纳米量子点 可检测与***癌成高度相关性的 δ-catenin蛋白的变化，从而用于诊断前 列腺癌。
虽然本发明已以较佳实施例披露如上， 然其并非用以限定本发明。 任 何人或动物的组织和各种体液中，只要存在可用 δ-catenin抗体或 δ-catenin 抗体与纳米量子点结合体识别的 δ-catenin蛋白， 则 δ-catenin抗体结合酶 联或结合纳米量子点便可用于检测。 任何所属技术领域的技术人员， 在不 脱离本发明的精神和范围内， 当可作些许的更动与改进， 因此本发明的保 护范围当视权利要求所界定者为准。

Claims

权 利 要 求 书
1. 一种检测或诊断***癌的试剂盒， 其特征在于包含 δ-catenin吸 附抗体、 包被缓冲液、 洗涤缓冲液、 封闭缓冲液、 δ-catenin检测抗体、 酶 标抗体。
2. 一种检测或诊断***癌的试剂盒， 其特征在于包含 δ-catenin吸 附抗体、 包被缓冲液、 洗涤缓冲液、 封闭缓冲液、 δ-catenin检测抗体、 纳 米量子点标记。
3. 根据权利要求 1或 2所述的试剂盒，其特征在于，还包含标准品和 阴性对照。
4. 根据权利要求 3 所述的试剂盒， 其特征在于， 所述标准品为人 δ-catenin标准蛋白。
5. 根据权利要求 1或 2所述的试剂盒， 其特征在于， 所述 δ-catenin 吸附抗体为 δ-catenin抗体。
6. 根据权利要求 1或 2所述的试剂盒，其特征在于，所述包被缓冲液 为碳酸盐或 dPBS。
7. 根据权利要求 1或 2所述的试剂盒，其特征在于，所述洗涤缓冲液 为含洗涤剂的 dPBS。
8. 根据权利要求 1或 2所述的试剂盒， 其特征在于， 所述 δ-catenin 检测抗体和 δ-catenin吸附抗体识别不同的 δ-catenin蛋白序列。
9. δ-catenin蛋白在制备检测或诊断***癌的制剂中的应用。
PCT/CN2013/000118 2012-02-14 2013-02-04 一种检测或诊断***癌的试剂盒 WO2013120394A1 (zh)

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