Alkaline Phosphatase Stain Solution (Coupling azo-dye Method)

堿性磷酸酶染色液（偶氮偶聯法）

産品信息

【溫馨提示】：見(jiàn)我司懋康生(shēng)物(wù)整理的堿性磷酸酶（Alkaline phosphatase，ALP，AKP）産品專題。

【溫馨提示】：見(jiàn)我司懋康生(shēng)物(wù)整理的堿性磷酸酶活性檢測試劑盒（定量檢測）産品專題。

【溫馨提示】：見(jiàn)我司懋康生(shēng)物(wù)整理的堿性磷酸酶染色檢測試劑盒（定性，幹細胞檢測）産品專題。

産品描述

堿性磷酸酶（Alkaline phosphatase，ALP，AKP或AP）是一(yī)類磷酸酯酶，堿性條件(jiàn)下(xià)能(néng)夠将核苷酸、蛋白(bái)、甾醇等分子去磷酸化，廣泛分布于哺乳動物(wù)組織内。ALP主要存在于物(wù)質交換活躍處（細胞膜），如腸上(shàng)皮和腎近曲小(xiǎo)管的刷狀緣、附睾上(shàng)皮之靜(jìng)纖毛、肝的毛細膽管膜以及微動脈和毛細血管動脈部之内皮。還(hái)發現于内質網、高(gāo)爾基複合體、吞飲小(xiǎo)泡、腸上(shàng)皮之溶酶體、中性粒細胞之中性顆粒以及平滑肌之細胞膜及吞飲小(xiǎo)泡。

本堿性磷酸酶染色液（偶氮偶聯法）（Alkaline Phosphatase Stain Solution (Coupling azo-dye Mehod)）利用的是偶氮偶聯法來定性檢測磷酸酶活性。工(gōng)作原理是在pH 9.2-9.8的堿性條件(jiàn)下(xià)，細胞内ALP可使AB-BI磷酸鹽水(shuǐ)解，釋放(fàng)出磷酸與萘酚，後者與偶聯重氮鹽生(shēng)成有色産物(wù)，定位于細胞質中。該染液可用于血液、骨髓或細胞塗片、冰凍切片、石蠟切片等的ALP染色，ALP陽性反應部位呈藍色，位于胞漿，染色結果比金屬鹽沉澱法相(xiàng)對可靠些。

産品組成

保存與運輸方法

保存：按照(zhào)對應組分的要求來保存，1年(nián)有效。

運輸：冰袋運輸。

注意事(shì)項

1）   血液、骨髓細胞塗片或其他樣本均應新鮮，薄厚适宜，及時固定，否則會(huì)影響酶的活性。

2）   培養細胞染色操作過程中，清洗、染色等步驟都應輕微，以免損傷或丢失細胞。

3）   培養細胞或爬片細胞不建議采用ALP固定液，此ALP固定液适用于血液或骨髓細胞塗片的固定。

4）   為(wèi)了您的安全和健康，請穿實驗服并戴一(yī)次性手套操作。

操作步驟

一(yī)、 ALP染色液的準備

臨用前，取适量的AS-BI染色液（B1）和FBB染色液（B2）按照(zhào)1：1比例混合，即得到(dào)ALP染色液，現配現用。

二、 塗片或切片染色

1.1    血液、骨髓或細胞塗片、冰凍切片、石蠟切片放(fàng)入ALP固定液固定3min（注：梯度入水(shuǐ)的石蠟切片無需固定），水(shuǐ)洗。

1.2    滴加新鮮配制的ALP染色液，放(fàng)入濕盒，避光(guāng)孵育15-20min，水(shuǐ)洗。

1.3    入核固紅(hóng)染色液複染3-5min。

1.4    水(shuǐ)洗、鏡檢或甘油明膠封片後鏡檢。

三、 貼壁細胞染色

2.1 取6孔闆或其他容器(qì)培養的細胞，棄培養液，PBS清洗細胞。

2.2 加入4%多(duō)聚甲醛固定10-20min，PBS清洗細胞。

2.3 滴加新鮮配制的ALP染色液，放(fàng)入濕盒，避光(guāng)孵育15-20min，PBS清洗。

2.4 入核固紅(hóng)染色液複染30s~3min。

2.5    PBS清洗、鏡檢。

三、染色結果

相(xiàng)關産品

文獻引用：

1）Fu, L., Jin, P., Hu, Y., Lu, H., & Su, L. (2020). KR‑12‑a6 promotes the osteogenic differentiation of human bone marrow mesenchymal stem cells via BMP/SMAD signaling. Molecular Medicine Reports, 21, 61-68.

For ALP staining, hBMSCs in each group were washedtwice with PBS at the end of the 7‑day KR‑12‑a6‑treatmentperiod, fixed with 4% paraformaldehyde for 30 s, and then stained using the ALP staining kit (Maokang Biotechnology, Shanghai)according to the manufacturer's instructions. images were acquired by an inverted phase contrast microscope (CKX41, Olympus) under x100 magnification. At leastfive fields per sample were randomly selected and observed.

1）Fu, L., Jin, P., Hu, Y., Lu, H., & Su, L. (2020). KR‑12‑a6 promotes the osteogenic differentiation of human bone marrow mesenchymal stem cells via BMP/SMAD signaling. Molecular Medicine Reports, 21, 61-68.

For ALP staining, hBMSCs in each group were washedtwice with PBS at the end of the 7‑day KR‑12‑a6‑treatmentperiod, fixed with 4% paraformaldehyde for 30 s, and then stained using the ALP staining kit (Maokang Biotechnology, Shanghai)according to the manufacturer's instructions. images were acquired by an inverted phase contrast microscope (CKX41, Olympus) under x100 magnification. At leastfive fields per sample were randomly selected and observed.

2)Peiying Lv, Pengfei Gao, Yanyan Yang, Zihui Wang, Lu Sun, Feifei Mo, Guangjie Tian, Mingjie Kuang, Yonglan Wang.Osteocyte-derived exosomes induced by mechanical strain promote human periodontal ligament stem cell proliferation and osteogenic differentiation in an inflammatory environment via the miR-181b-5p/PTEN/AKT signaling pathway.Stem Cell Research & Therapy；

The activity of alkaline phosphatase (ALP) was analyzed using an Alkaline Phosphatase Staining Kit (MKbio, Shanghai, China). Staining images were acquired using an inverted microscope, and ImageJ was used to analyze the images.

 — —Written/Edited by V. Shallan【版權歸MKBio懋康所有】

上海夢澤生物科技有限公司是一(yī)家涉足于生(shēng)命科學和生(shēng)物(wù)技(jì)術(shù)領域研究的試劑、儀器(qì)和實驗室消耗品與實驗服務工(gōng)作，主要從(cóng)事(shì)細胞生(shēng)物(wù)學、植物(wù)學、分子生(shēng)物(wù)學、免疫學、生(shēng)物(wù)化學、蛋白(bái)組學。生(shēng)物(wù)制藥與診斷試劑研發生(shēng)産等領域。 本公司秉承“以人為(wèi)本，以誠為(wèi)信、合同守信”的經營理念。堅持"品質保障"的原則為(wèi)廣大客戶提供優質産品。