FITC膜聯(lián)蛋白V細(xì)胞凋亡檢測(cè)試劑盒（BD Pharmingen）描述

凋亡是在胚胎發(fā)育過(guò)程中發(fā)生的正常生理過(guò)程以及組織穩(wěn)態(tài)的維持。凋亡程序的特征是某些形態(tài)學(xué)特征，包括質(zhì)膜不對(duì)稱(chēng)和附著，細(xì)胞質(zhì)和細(xì)胞核的縮合以及DNA的核小體間裂解。質(zhì)膜的損失是zui早的特征之一。在凋亡細(xì)胞中，膜磷脂磷脂酰絲氨酸（PS）從質(zhì)膜的內(nèi)部和外部葉片轉(zhuǎn)位，從而將PS暴露于外部細(xì)胞環(huán)境。膜聯(lián)蛋白V是一種35-36 kDa的Ca2 +依賴(lài)性磷脂結(jié)合蛋白，對(duì)PS具有高親和力，并與暴露的PS結(jié)合細(xì)胞。膜聯(lián)蛋白V可以與包括FITC的熒光染料綴合。該格式保留了對(duì)PS的高親和力，因此用作對(duì)正在進(jìn)行細(xì)胞凋亡的細(xì)胞的流式細(xì)胞分析的敏感探針。由于PS的外化發(fā)生在凋亡的早期階段，F(xiàn)ITC膜聯(lián)蛋白V染色可以比基于核變化如DNA片段化的測(cè)定在早期鑒定凋亡。

FITC膜聯(lián)蛋白V染色先于膜完整性的喪失，伴隨著由凋亡或壞死過(guò)程引起的細(xì)胞死亡階段。因此，F(xiàn)ITC膜聯(lián)蛋白V的染色通常與重要染料（如碘化丙啶（PI）或7-氨基放線(xiàn)菌素（7-AAD））結(jié)合使用，以使研究人員能夠鑒定早期凋亡細(xì)胞（PI陰性，F(xiàn)ITC膜聯(lián)蛋白V正）。具有完整膜的活細(xì)胞排除PI，使死亡和受損細(xì)胞的膜可透過(guò)PI。例如，認(rèn)為可行的細(xì)胞是FITC膜聯(lián)蛋白V和PI陰性; 早期凋亡細(xì)胞是FITC Annexin V陽(yáng)性和PI陰性; 細(xì)胞凋亡或已經(jīng)死亡的細(xì)胞都是FITC膜聯(lián)蛋白V和PI陽(yáng)性。該測(cè)定不區(qū)分已經(jīng)經(jīng)歷凋亡死亡的細(xì)胞與由于壞死性途徑死亡的細(xì)胞，因?yàn)樵谌我磺闆r下，死細(xì)胞都會(huì)用FITC Annexin V和PI染色。然而，隨著時(shí)間的推移，當(dāng)細(xì)胞凋亡被測(cè)量時(shí)，細(xì)胞可以經(jīng)常從FITC膜聯(lián)蛋白V和PI陰性（可行或無(wú)可測(cè)量的凋亡）追溯到FITC膜聯(lián)蛋白V陽(yáng)性和PI陰性（早期凋亡，膜完整性存在），zui后到FITC膜聯(lián)蛋白V和PI陽(yáng)性（末期凋亡和死亡）。細(xì)胞通過(guò)這三個(gè)階段的運(yùn)動(dòng)表明細(xì)胞凋亡。相比之下，一個(gè)單一的觀察結(jié)果表明，細(xì)胞都是FITC膜聯(lián)蛋白V和PI陽(yáng)性，本身就顯示出關(guān)于細(xì)胞進(jìn)行死亡的過(guò)程的信息較少。

準(zhǔn)備和儲(chǔ)存

在4°C下未經(jīng)稀釋保存，防止長(zhǎng)時(shí)間暴露于光線(xiàn)下。不要凍結(jié)

產(chǎn)品通知

1. 由于應(yīng)用程序不同，每個(gè)研究者應(yīng)滴定試劑以獲得*結(jié)果。
2. 所有血清蛋白的來(lái)源均來(lái)自美國(guó)農(nóng)業(yè)部檢查的位于美國(guó)的屠宰場(chǎng)。
3. 注意：疊氮鈉在酸性條件下會(huì)產(chǎn)生高毒性的疊氮酸。在棄置之前將水中的疊氮化合物稀釋?zhuān)员苊庠诠艿乐欣鄯e潛在的爆炸性沉積物。

DESCRIPTION

Apoptosis is a normal physiologic process which occurs during embryonic development as well as in maintenence of tissue homeostasis. The apoptotic program is characterized by certain morphologic features, including loss of plasma membrane asymmetry and attachment, condensation of the cytoplasm and nucleus, and internucleosomal cleavage of DNA. Loss of plasma membrane is one of the earliest features. In apoptotic cells, the membrane phospholipid phosphatidylserine (PS) is translocated from the inner to the outer leaflet of the plasma membrane, thereby exposing PS to the external cellular environment. Annexin V is a 35-36 kDa Ca2+ dependent phospholipid-binding protein that has a high affinity for PS, and binds to cells with exposed PS. Annexin V may be conjugated to fluorochromes including FITC. This format retains its high affinity for PS and thus serves as a sensitive probe for flow cytometric analysis of cells that are undergoing apoptosis. Since externalization of PS occurs in the earlier stages of apoptosis, FITC Annexin V staining can identify apoptosis at an earlier stage than assays based on nuclear changes such as DNA fragmentation.

FITC Annexin V staining precedes the loss of membrane integrity which accompanies the latest stages of cell death resulting from either apoptotic or necrotic processes. Therefore, staining with FITC Annexin V is typically used in conjunction with a vital dye such as propidium iodide (PI) or 7-Amino-Actinomycin (7-AAD) to allow the investigator to identify early apoptotic cells (PI negative, FITC Annexin V positive). Viable cells with intact membranes exclude PI, wheras the membranes of dead and damaged cells are permeable to PI. For example, cells that are considered viable are FITC Annexin V and PI negative; cells that are in early apoptosis are FITC Annexin V positive and PI negative; and cells that are in late apoptosis or already dead are are both FITC Annexin V and PI positive. This assay does not distinguish between cells that have undergone apoptotic death versus those that have died as a result of a necrotic pathway because in either case, the dead cells will stain with both FITC Annexin V and PI. However, when apoptosis is measured over time, cells can be often tracked from FITC Annexin V and PI negative (viable, or no measurable apoptosis), to FITC Annexin V positive and PI negative (early apoptosis, membrane integrity is present) and finally to FITC Annexin V and PI positive (end stage apoptosis and death). The movement of cells through these three stages suggests apoptosis. In contrast, a single observation indicating that cells are both FITC Annexin V and PI positive, in of itself, reveals less information about the process by which the cells underwent their demise.

FITC膜聯(lián)蛋白V細(xì)胞凋亡檢測(cè)試劑盒（BD Pharmingen）PREPARATION AND STORAGE

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.

PRODUCT NOTICES

1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
2. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
3. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.