| 產(chǎn)品編號 | bs-8004-10 |
| 英文名稱(chēng) | TUNEL |
| 中文名稱(chēng) | TUNEL細胞凋亡檢測試劑盒 |
| 別 名 | TUNEL Apoptosis Assay Kit; In Situ Cell Death Detection Kit; In Situ Cell Death Detection Kit, POD; TUNEL細胞凋亡檢測試劑盒(熒光法); 細胞凋亡檢測試劑盒;細胞凋亡原位檢測試劑盒; 細胞凋亡試劑盒; |
| 克 隆 號 | |
| CAS | |
| 理論分子量 | kDa |
| 檢測分子量 | |
| 保存條件 | Store at -15 to -25℃. |
| 產(chǎn)品介紹 |
Widely used methods to determine apoptosis include the analysis of the genomic DNA by agarose-gel electrophoresis and DNA fragmentation assays based on 3H-thymidine and, alternatively, 5-Bromo-2'-deoxy-uridine. The methods involve the separation of fragmented, low molecular-weight DNA from unfragmented, high molecular-weight DNA in a given cell population. Thus, these methods do not provide information about the fate of an individual cell in a given cell population or, particularly, in tissue sections. Alternatively, individual apoptotic cells may be microscopically recognized because of the characteristic appearance of nuclear chromatin condensation and fragmentation, but this method is subjective and limited to a relatively narrow time window when the morphological changes are at a maximum. The hallmark of apoptosis is DNA degradation, which in early stages is selective to the internucleosomal DNA linker regions. The DNA cleavage may yield double-stranded and single-stranded DNA breaks (nicks). Both types of breaks can be detected by labeling the free 3'-OH termini with modified nucleotides (e.g., biotin-dUTP, DIG-dUTP, fluorescein-dUTP) in an enzymatic reaction. The enzyme terminal deoxynucleotidyl transferase (TdT) catalyzes the template-independent polymerization of deoxyribonucleotides to the 3'-end of single- and double-stranded DNA. This method has also been termed TUNEL (TdT-mediated dUTP-X nick end labeling). Alternatively, free 3'-OH groups may be labeled using DNA polymerases by the template-dependent mechanism called nick translation. However, the TUNEL method is considered to be more sensitive and faster. Contents: 1. Enzyme Solution (TdT), 5 vials 2. Label Solution (fluorescein-dUTP), 5 vials 3. Converter POD (anti-fluorescein antibody-POD), ready-to-use Principle The In Situ Cell Death Detection Kit, POD is based on the detection of single- and double-stranded DNA breaks that occur at the early stages of apoptosis. Apoptotic cells are fixed and permeabilized. Subsequently, the cells are incubated with the TUNEL reaction mixture that contains TdT and fluorescein-dUTP. During this incubation period, TdT catalyzes the addition of fluorescein-dUTP at free 3'-OH groups in single- and double-stranded DNA. After washing, the label incorporated at the damaged sites of the DNA is marked by an anti-fluorescein antibody conjugated with the reporter enzyme peroxidase. After washing to remove unbound enzyme conjugate, the POD retained in the immune complex is visualized by a substrate reaction. 凋亡是許多正常的生理過(guò)程所必需的，包括免疫系統的成熟和作用機制、組織器官和肢體的發(fā)生、神經(jīng)系統的發(fā)生等過(guò)程。在許多病理條件下，凋亡機制的調節失衡起很重要的作用，包括組織細胞的缺血、缺氧、出血、中風(fēng)、心臟病、癌癥、艾滋病、自身免疫缺損和中樞神經(jīng)系統的退行性變等。在癌基因的研究方面，由于細胞凋亡可由抗癌藥物、放射及高熱等啟動(dòng)，并且腫瘤細胞具有由于一些癌基因表達即能啟動(dòng)凋亡的內在特性，因此細胞凋亡已引起人們的廣泛重視。 檢測凋亡細胞的方法有數種。細胞凋亡過(guò)程中核酸內切酶的活化是關(guān)鍵的過(guò)程，導致核DNA被裂解成寡核苷酸大小的片段。因此，這個(gè)過(guò)程被用于檢測凋亡，在瓊脂糖凝膠電泳上出現典型的“DNA梯形帶”。但是，這種方法既不能檢測單個(gè)細胞水平的凋亡，又不能提供細胞發(fā)生凋亡所處的組織位置和細胞分化狀態(tài)等方面的信息。 這一切可以由凋亡的原位酶標記方法來(lái)完成。DNA聚合酶和末端脫氧核糖核酸轉移酶（TdT）用來(lái)標記DNA鏈斷端的核酸，應用TdT進(jìn)行的末端反應又稱(chēng)為ISEL或TUNEL技術(shù)，它與應用DNA聚合酶的ISNT（in situ nick translation）相比具有以下優(yōu)點(diǎn)：靈敏度高、快速、優(yōu)先標記發(fā)生凋亡的細胞，而不是壞死的細胞。 本試劑盒檢測細胞凋亡分以下三個(gè)步驟：1、標記DNA鏈斷端，由TdT催化，以FITC標記的核苷酸為原料，進(jìn)行非模板依賴(lài)方式的3?OH端DNA末端聚合反應；2、用HRP或AP標記的羊抗FITC的抗體（Fab段）與聚合到DNA末端的FITC結合；3、用酶的底物顯色，HRP常用DAB顯色，AP常用BCIP/NBT顯色。 細胞凋亡原位檢測:我公司提供代測服務(wù)： 1.只根據檢測樣本數量訂購試劑盒即可，所有輔助試劑免費； 2.可提供組織標本-石蠟包埋好的蠟塊或切好的片子快遞至我公司即可，每做一張片子收取60.00元的服務(wù)費用； |
