总膜和质膜蛋白分离试剂盒

Minute^TM 膜蛋白提取试剂盒

目录号：SM-005

描述  

      Minute 膜蛋白提取试剂盒通过离心管柱专利技术配合特定裂解液可以提取天然的膜蛋白。其原理是细胞/组织通过Buffer A致敏，然后细胞以Z字形路径通过离心管柱。在这个过程中细胞膜会破裂，分离出完整的细胞核，因此最终获得的膜蛋白中不含有核膜和核蛋白污染。通过差速离心和密度离心可以把细胞蛋白分为：总膜，细胞核，细胞质，细胞器及质膜，而无需超高速离心。不同于其他试剂盒，因为高速离心通过离心管柱时使细胞膜破裂，所以不需要匀浆，匀浆通常会引起膜蛋白提取结果的差异。而我们的试剂盒使用同样起始量的细胞，只需设定离心力和时间，结果一致性更好。整个操作过程不超过45分钟即可完成。

应用  

本试剂盒可以快速从细胞或组织中分离天然的膜蛋白，可应用于SDS-PAGE, immunoblottings, ELISA, IP,膜蛋白结构分析, 2-D,酶活性检测等其他应用。

 

 Figure 1. A. SDS-PAGE profiles of total cell lysate (TC) and isolated plasma membrane proteins (PM) from human and rat cultured cells. Lane 1, Human lung cancer cell A549 total cell lysate; Lane 2, Isolated PM of A549; Lane 3, PM of rat REL-6TN cells; Lane 4, Total cell lysate of rat REL-6TN cells. 
B. Western blottings : Proteins in A were transferred to nitrocellulose membrane and probed with anti-Na/K ATPase alpha1, a plasma membrane marker (Upstate, Clone 464.6)) and anti-lamin B1, a nuclear envelope marker (ab16048, abcam Cambridge, MA). The specific protein bands were visualized by a color metric substrate Opti-4CN (Bio-RAD). Figure 1. A. SDS-PAGE profiles of total cell lysate (TC) and isolated plasma membrane proteins (PM) from human and rat cultured cells. Lane 1, Human lung cancer cell A549 total cell lysate; Lane 2, Isolated PM of A549; Lane 3, PM of rat REL-6TN cells; Lane 4, Total cell lysate of rat REL-6TN cells. 
B. Western blottings : Proteins in A were transferred to nitrocellulose membrane and probed with anti-Na/K ATPase alpha1, a plasma membrane marker (Upstate, Clone 464.6)) and anti-lamin B1, a nuclear envelope marker (ab16048, abcam Cambridge, MA). The specific protein bands were visualized by a color metric substrate Opti-4CN (Bio-RAD). 
C. Densitometry measurement of Na/K ATPase alpha1 signals in B (TC vs. PM). Total cell lysates were extracted by Minute Denaturing Total Protein Extraction Kit (SD-001, Invent Biotechnologies, Eden Prairie, MN).
C. Densitometry measurement of Na/K ATPase alpha1 signals in B (TC vs. PM). Total cell lysates were extracted by Minute Denaturing Total Protein Extraction Kit (SD-001, Invent Biotechnologies, Eden Prairie, MN).  Figure 2 A. SDS-PAGE profiles of isolated total membrane proteins from mouse tissues (T=Total Cell lysate, C=Cytosol Fraction, M= Total Membrane Fraction) 
B. Proteins shown in A were transferred to a nitrocellulose membrane and probed with rabbit-anti mouse pan-cadherin (ab6529,abcam, Cambridge, MA), and anti-actin by Western blotting. The specific protein bands were visualized by a color metric substrate Opti-4CN (Bio-RAD). Signals of pan-cadherin (a 130 kda plasma membrane marker) were significantly enhanced in total membrane protein fractions. Total cell lysates were extracted by MinuteTM Denaturing Total Protein Extraction Kit (SD-001, Invent Biotechnologies, Eden Prairie, MN). 

 

Figure 3. A. SDS-PAGE profiles of total membrane protein fraction (TM) and isolated plasma membrane protein fraction from mouse tissues. 
B. Western blotting of proteins in A were transferred to nitrocellulose membrane and probed with rabbit anti-cadherin (abcam, Cambridge, MA). The specific protein bands were visualized by a color metric substrate Opti-4CN (Bio-RAD). 
C. Densitometry measurement of cadherin signals in B (TM vs. PM).