This Human Myocilin overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Myocilin protein (Cat: 13090-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the full length of human MYOC (Q99972) (Met 1-Met 504) was fused with a polyhistidine tag at the C-terminus.
The secreted recombinant human MYOC consists of 483 amino acids and has a predicted molecular mass of 54.7 kDa. Since most of the MYOC was cleaved at Glu 214, generating a C-terminal fragment of 32 kDa, the purified human MYOC migrates as an approximately 33 kDa band in SDS-PAGE under reducing conditions.
Human Myocilin HEK293 Overexpression Lysate: 使用指南
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
稳定性 & 储存条件
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human Myocilin HEK293 Overexpression Lysate: 别称
Human GLC1A Overexpression Lysate; Human GPOA Overexpression Lysate; Human JOAG Overexpression Lysate; Human JOAG1 Overexpression Lysate; Human myocilin Overexpression Lysate; Human TIGR Overexpression Lysate
Myocilin, also known as Trabecular meshwork-induced glucocorticoid response protein, MYOC, and GLC1A, is a protein that contains one olfactomedin-like domain. Myocilin / MYOC may participate in the obstruction of fluid outflow in the trabecular meshwork. Myocilin / MYOC is expressed in large amounts in various types of muscle, ciliary body, papillary sphincter, skeletal muscle, heart, and other tissues. Myocilin / MYOC is expressed predominantly in the retina. In normal eyes, it is found in the inner uveal meshwork region and the anterior portion of the meshwork. In contrast, in many glaucomatous eyes, it is found in more regions of the meshwork and appeared more intensively than in normal eyes, regardless of the type of clinical severity of glaucoma. Defects in Myocilin / MYOC may contribute to primary congenital glaucoma type 3A (GLC3A). Defects in MYOC may also contribute to this phenotype via digenic inheritance. GLC3A is an autosomal recessive form of primary congenital glaucoma (PCG). PCG is characterized by a marked increase of intraocular pressure at birth or early childhood, large ocular globes (buphthalmos), and corneal edema.