Human ARSA HEK293 Overexpression Lysate

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Human ARSA HEK293 Overexpression Lysate: 製品の情報

製品の説明
This Human ARSA overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of ARSA protein (Cat: 10449-H08H) from the overexpression lysate was verified.
発現ホスト
HEK293 Cells
Human
タンパク質構築情報
A DNA sequence encoding the human Arylsulfatase A (NP_000478.2) (Met 1-Ala 507) was expressed with a C-terminal polyhistidine tag.
分子量
The recombinant human ARSA consists of 500 amino acids and predicts a molecular mass of 53 kDa. as estimated by SDS-PAGE under reducing conditions.

Human ARSA HEK293 Overexpression Lysate: 用法

調製方法
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.
溶解バッファー
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
おすすめの用法
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.
バッファー
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
安定性 & 保存条件
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.
アプリケーション
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human ARSA HEK293 Overexpression Lysate: 別名

Human MLD Overexpression Lysate

ARSA 背景情報

Arylsulfatase A (ARSA) is synthesized as a 52KDa lysosomal enzyme. It is a member of the sulfatase family that is required for the lysosomal degradation of cerebroside-3-sulfate, a sphingolipid sulfate ester and a major constituent of the myelin sheet. Arylsulfatase A is activated by a required co- or posttranslational modification with the oxidation of cysteine to formylglycine. Metachromatic leukodystrophy (MLD) is a lysosomal storage disease in the central and peripheral nervous systems with severe and progressive neurological symptoms caused by the deficiency of Arylsulfatase A. Deficiency of this enzyme is also found in apparently healthy individuals, a condition for which the term pseudodeficiency is introduced. ARSA forms dimers after receiving three N-linked oligosaccharides in the endoplasmic reticulum, and then the dimers are transported to the Golgi where they receive mannose 6-phosphate recognition markers. And thus, ARSA is transported and delivered to dense lysosomes in a mannose 6-phosphate receptor-dependent manner. It has been shown that within the lysosomes, the ARSA dimers can oligomerize to an octamer in a pH-dependent manner. The ARSA deficiency leads to metachromatic leucodystrophy (MLD), a lysosomal storage disorder associated with severe and progressive demyelination in he central and peripheral nervous system. Additionally, the serum level of arylsulfatase A might be helpful in diagnosis of lung and central nervous system cancer.
完全な名称
arylsulfatase A
参考文献
  • Laidler PM. (1991) Arylsulfatase A--physico-chemical properties and the use of enzyme radioimmunoassay in medical diagnosis Folia Med Cracov. 32(3-4): 149-68.
  • Jean S, et al. (2006) Ethanol decreases rat hepatic arylsulfatase A activity levels. Alcohol Clin Exp Res. 30(11): 1950-5.
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