Certificate of Analysis and Data Sheet

Source:
HEK293
(Human embryonic kidney cell line)

Catalog No.
CYT-247

Introduction:

Adiponectin (also called ACRP30, GBP28 and apM1) is a hormone exclusively expressed from adipose tissue 1. Many studies demonstrate that adiponectin has direct anti-diabetic, anti-atherogenic and anti-inflammatory functions. Adiponectin can increase insulin sensitivity of skeletal muscle 2, attenuate hepatic lipogenesis 3 and luconeogenesis 4, regulate NO production in endothelial cells 5, inhibit proliefeartion of smooth muscle cells 6 and prevent lipid accumulation of macrophage cells 7.
In the circulation, adiponectin is present as three different oligomeric complexes, including the high molecular weight (HMW), the middle molecular weight (MMW, also called hexamer) and low molecular weigh (MMW, also called trimer) forms 8. Different oligomeric complex of adiponectin activates different signaling pathways and exerts distinct functions.
Trimeric form of adiponectin was expressed in HEK293 cells stably transfected with pcmAd-F vector encoding FLAG epitope-tagged mouse adiponectin. Recombinant mouse adiponectin ?Trimeric form was purified using anti-FLAG M2 affinity gel (Sigma-Aldrich) and was eluted with FLAG peptide. FLAG peptides were removed by an extensive wash with an excessive volume of saline. Highly sensitive matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass-spectrometric analysis confirmed that FLAG peptide was not detectable in the protein solution.

Specificity:

The cystenine 39 was replaced with alanine (C39A) 9. mAd-C39A can only form trimer, but not hexamer or HMW form.

Purity:

>95%

Activity:

ED50= 3-8.5 μg/mL, as determined by its ability to inhibit proliferation of HASMCs induced by HB EGF.

Formulation:

Sterile filtered and lyophilized from 0.5 mg/ml in 0.05 M phosphate buffer, 0.05 M NaCl, pH 7.2.

Reconstitution:

Add 0.2 ml of deionized water and let the lyophilized pellet dissolve completely.

Storage:

Store lyophilized protein at -20蚓. Aliquot the product after reconstitution to avoid repeated freezing/thawing cycles. Reconstituted protein can be stored at 4蚓 for a limited period of time; it does not show any change after two weeks at 4蚓.

Stability/Shelf Life:

The lyophilized protein remains stable until the expiry date when stored at -20蚓.

Applications:

In vitro, ex vivo and in vivo activity analysis, binding assay et al.

References:

1. Wang Y, Lam KS, Xu JY, Lu G, Xu LY, Cooper GJ, Xu A. Adiponectin inhibits cell proliferation by interacting with several growth factors in an oligomerization-dependent manner. J Biol Chem. 2005 May 6;280(18):18341-7.
2. Berg AH, Combs TP, Scherer PE. ACRP30/adiponectin: an adipokine regulating glucose and lipid metabolism. Trends Endocrinol Metab. 2002;13:84-89.
3. Yamauchi T, Kamon J, et al. The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity. Nat Med. 2001;7:941-946.
4. Xu A, Wang Y, Keshaw H, Xu LY, Lam KS, Cooper GJ. The fat-derived hormone adiponectin alleviates alcoholic and nonalcoholic fatty liver diseases in mice. J Clin Invest. 2003;112:91- 100.
5. Berg AH, Combs TP, Du X, Brownlee M, Scherer PE. The adipocyte-secreted protein Acrp30 enhances hepatic insulin action. Nat Med. 2001;7:947-953.
6. Chen H, Montagnani M, Funahashi T, Shimomura I, Quon MJ. Adiponectin stimulates production of nitric oxide in vascular endothelial cells. J Biol Chem. 2003;278:45021-45026.
7. Matsuda M, Shimomura I, et al. Role of adiponectin in preventing vascular stenosis. The missing link of adipo-vascular axis. J Biol Chem. 2002;277:37487-37491.
8. Okamoto Y, Kihara S, et al. Adiponectin reduces atherosclerosis in apolipoprotein E-deficient mice. Circulation. 2002;106:2767-2770.
9. Kobayashi H, Ouchi N, Kihara S, Walsh K, Kumada M, Abe Y, Funahashi T, Matsuzawa Y, Ferre P. Selective Suppression of Endothelial Cell Apoptosis by the High Molecular Weight.

Usage:

Prospec's products are furnished for LABORATORY RESEARCH USE ONLY. The product may not be used as drugs, agricultural or pesticidal products, food additives or household chemicals.

Latest Publications:

1. A longitudinal study of serum adiponectin during normal pregnancy.
BJOG 2006 Jan;113(1):110-3
2. High molecular weight adiponectin correlates with insulin sensitivity in patients with hepatitis C genotype 3, but not genotype 1 infection.
Am J Gastroenterol 2005 Dec;100(12):2717-23
3. Single Nucleotide Polymorphisms in exon 3 of the adiponectin gene in subjects with type 2 diabetes mellitus.
Rocz Akad Med Bialymst 2005;50:148-50
4. Decreased serum adiponectin: an early event in pediatric nonalcoholic fatty liver disease.
J Pediatr 2005 Dec;147(6):835-8
5. Endothelial cell IL-8, a new target for adiponectin: implications in vascular protection.
Circ Res 2005 Dec 9;97(12):1216-9
6. Relationship between plasma adiponectin levels and metabolic risk profiles in Taiwanese children.
Obes Res 2005 Nov;13(11):2014-20