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人胎球蛋白A（Fetuin-A）酶免试剂盒
美国*
【型号】 KT-800
【预期用途】
这个ELISA试剂盒是用来对血清、血浆、细胞培养上清液、组织提取液以及尿液中的人胎球蛋白A也叫做α-2-Heremans-Schmid糖蛋白（α2–Heremans- Schmid glycoprotein, AHSG）的浓度进行定量检测。可以通过对胎球蛋白A的检测来诊断某些癌症和血清蛋白的基因遗传缺陷。血清胎球蛋白A下降是体内炎症的重要指标，并与冠状动脉硬化和心脏钙化密切相关；尿液胎球蛋白A与肾功能相关。该胎球蛋白酶免试剂盒仅供实验研究使用。
【生理学简介】
胎球蛋白A也叫做α-2-Heremans-Schmid糖蛋白，它的分子量是59kDa，拥有两个氨基末端半胱胺酸蛋白酶抑制因子和一个较小的羧基末端区域。胎球蛋白A通过肝脏合成，分泌到血液中。一般来说，成人血液中胎球蛋白A的浓度范围为0.5-1.0g/L。在胎儿体内，血清中胎球蛋白A呈现出高浓度。它会参与蛋白酶抑制活动以及钙代谢和骨生成的相关活动。胎球蛋白A在骨骼和牙齿中积累，是生物学上的非胶原骨蛋白成分的主要部分。经研究证明，胎球蛋白A会阻止钙沉淀，是循环过程中的zui主要的钙化抑制剂。zui近的研究表明，尿毒症、血液透析患者的胎球蛋白A的浓度比正常健康人的低。肾功能衰竭患者的胎球蛋白A浓度低与心血管死亡率增高相关。另一方面，老年血清胎球蛋白A增高，更易患糖尿病，它是一个*的胰岛素灵敏度下降指标。甚至，有科研结果表明高胎球蛋白A浓度可以作为心血管疾病患者的一个独立指标。
【检验原理】
这个ELISA的设计，研发和生产的目的是对血清中人胎球蛋白A进行定量检验。该试剂盒是基于运用两种特定羊抗人胎球蛋白A多抗与人胎球蛋白A的不同抗原表位结合的“双位点夹心”技术。
将标准品、质控品、和含胎球蛋白A的预稀释的的病人血清样本加入到高度亲和性的羊抗人胎球蛋白A抗体包被的微孔板中。经过*个保温孵育期后，人胎球蛋白A被微孔板内壁的抗体吸附住，而各个孔中的游离蛋白质被冲走。随后，HRP标记的多抗抗人胎球蛋白A抗体加入到各个孔中，形成了“捕捉抗体-人胎球蛋白A-HRP标记示踪抗体”的“双位点夹心”结构。游离的示踪抗体在随后的清洗过程中被冲走。HRP标记的示踪抗体被吸附到微孔板的内壁，在3,3',5,5'-四甲基联苯胺（TMB）和过氧化氢的混合基质溶液中经过一段时间的反应，测量吸光度。微孔板内壁的胎球蛋白A与示踪抗体连结的酶活性与样本中胎球蛋白A的浓度直接成正比例。根据标准品中不同的胎球蛋白A的浓度的吸光度，按照“点到点”或者“体积比例”绘制标准曲线，待测样品中胎球蛋白A的浓度可以直接从标准曲线按测量到的吸光度得到。
【组成成分】
1. 胎球蛋白A抗体包被的微孔板：1块×96个
2. 胎球蛋白A示踪抗体：1瓶×0.6mL
3. 示踪抗体稀释液：1瓶×12mL
4. 胎球蛋白A检测浓缩缓冲液：1瓶×11mL
5. ELISA 浓缩洗涤液，30X ：1瓶×20mL
6. ELISA HRP基质：1瓶×12mL
7. ELISA终止液：1瓶×12mL
8. 胎球蛋白A标准品：5瓶
9. 胎球蛋白A质控品：2瓶
【应用本试剂盒已发表的文献】
1. Stenvinkel P, Wang K, Qureshi AR, Axelsson J, Pecoits-Filho R, Gao P, Barany P, Lindholm B, Jogestrand T, Heimbürger O, Holmes C, Schalling M, Nordfors L. Low fetuin-A levels are associated with cardiovascular death: Impact of variations in the gene encoding fetuin. Kidney Int. 2005 Jun;67（6）:2383-92.
2. Wang AY, Woo J, Lam CW, Wang M, Chan IH, Gao P, Lui SF, Li PK, Sanderson JE. Associations of serum fetuin-A with malnutrition, inflammation, atherosclerosis and valvular calcification syndrome and outcome in peritoneal dialysis patients. Nephrol Dial Transplant. 2005 Aug;20（8）:1676-85.
3. Sato H, Kazama JJ, Wada Y, Kuroda T, Narita I, Gejyo F, Gao P, Yamashita H. Decreased levels of circulating alpha2-Heremans-Schmid glycoprotein/Fetuin-A （AHSG） in patients with rheumatoid arthritis. Intern Med. 2007;46（20）:1685-91. Epub 2007 Oct 15.
4. Lehtinen AB, Burdon KP, Lewis JP, Langefeld CD, Ziegler JT, Rich SS, Register TC, Carr JJ, Freedman BI, Bowden DW. Association of alpha2-Heremans-Schmid glycoprotein polymorphisms with subclinical atherosclerosis. J Clin Endocrinol Metab. 2007 Jan;92（1）:345-52. Epub 2006 Oct 24
5. Lim P, Collet JP, Moutereau S, Guigui N, Mitchell-Heggs L, Loric S, Bernard M, Benhamed S, Montalescot G, Randé JL, Guéret P. Fetuin-A is an independent predictor of death after ST-elevation myocardial infarction. Clin Chem. 2007 Oct;53（10）:1835-40. Epub 2007 Aug 16
6. Ix JH, Wassel CL, Kanaya AM, Vittinghoff E, Johnson KC, Koster A, Cauley JA, Harris TB, Cummings SR, Shlipak MG; Health ABC Study. Fetuin-A and incident diabetes mellitus in older persons. JAMA. 2008 Jul 9;300（2）:182-8.
7. Ix JH, Wassel CL, Bauer DC, Toroian D, Tylavsky FA, Cauley JA, Harris TB, Price PA, Cummings SR, Shlipak MG; for the Health ABC Study. Fetuin-A and Bone Mineral Density in Older Persons: The Health Aging and Body Composition （Health ABC） Study. J Bone Miner Res. 2008 Nov 18.
8. Giulia Bivona, Chiara Bellia, Antonietta Caruso, Daniela Butera, Bruna Lo Sasso, Patrizia Altavilla, Rosa C. Carollo, Gaia Chiarello andMarcello Ciaccio. Low Serum Fetuin A Levels and Cardiovascular Events in End-Stage Renal Disease （ESRD） Patients. Research Journal of Medical Sciences 2 （4）: 200-202, 2008
9. Mario Cozzolinoa, Andrea Galassia, Maria Luisa Biondib, Olivia Turrib, Sergio Papagnic, Nicola Mongellic, Luigi Civitac, Maurizio Gallienia, Diego Brancaccioa Serum Fetuin-A Levels Link Inflammation and Cardiovascular Calcification in Hemodialysis Patients. Am J Nephrol 2006;26:423-429
10. G. Metry, P. Stenvinkel, A. R. Qureshi, J. J. Carrero, M. I. Yilmaz, P. Bárány, S. Snaedal, O. Heimbürger, B. Lindholm and M. E. Suliman Low serum fetuin-A concentration predicts poor outcome only in the presence of inflammation in prevalent haemodialysis patients. European Journal of Clinical Investigation 2008;38（11）:804 - 811
11. Doris Hendig, Veronika Schulz, Marius Arndt, Christiane Szliska, Knut Kleesiek, and Christian Go¨ tting1. Role of Serum Fetuin-A, a Major Inhibitor of Systemic Calcification, in Pseudoxanthoma Elasticum. Clin Chem 2006;52:227-234
12. G. Marhaug, V. Shah, R. Shroff, H. Varsani, L. R. Wedderburn, C. A. Pilkington and P. A. Brogan. Age-dependent inhibition of ectopic calcification: a possible role for fetuin-A and osteopontin in patients with juvenile dermatomyositis with calcinosis. Rheumatology 2008 47（7）:1031-1037
13. Marcello Ciaccio, et al. Changes in serum fetuin-A and inflammatory markers levels in end-stage renal disease （ESRD）: effect of a single session haemodialysis. Clin Chem Lab Med 2008;46:212–214
14. J. J. Carrero, P. Stenvinkel, B. Fellström, A. R. Qureshi, K. Lamb , O. Heimbürger, P. Bárány, K. Radhakrishnan, B. Lindholm, I. Soveri, L. Nordfors & P. G. Shiels. omere attrition is associated with inflammation, low fetuin-A levels and high mortality in prevalent haemodialysis patients. Journal of Internal Medicine 2007; 263（3）:302 – 312
15. C . Fiore , G . Celotta , G . Politi , L . Di Pino , Z . Casli , R . Mangiafico , S . Signorelli , P . Pennisi . Association of high alpha2-Heremans–Schmid glycoprotein/fetuin concentration in serum and intima-media thickness in patients with atherosclerotic vascular disease and low bone mass. Atherosclerosis 2007;195:110-115.
16. Sandro Mazzaferro, Marzia Pasquali, Francesco Pugliese, Giusi Barresi, Iacopo Carbone, Marco Francone, Daniela Sardella, Franco Taggi. Serum Levels of Calcification Inhibition Proteins and Coronary Artery Calcium Score: Comparison between Transplantation and Dialysis. Am J Nephrol 2007;27:75-83.
Human Fetuin-A ELISA Kit
Catalog Number: KT-800
This ELISA （enzyme-linked immunosorbent assay） kit is intended for the quantitative determination of human Fetuin-A, also known as alpha-2-HS glycoprotein （AHSG）, in serum, plasma, cell culture supernatant, tissue extraction and urine. The measurement of Fetuin-A in serum or plasma aids in the diagnosis of some cancers and genetically inherited deficiencies of this serum protein. This Fetuin-A ELISA kit is for laboratory professional use.
Fetuin-A, also known as alpha-2-HS glycoprotein, is a 59 kDa glycoprotein that consists of two amino-terminal cystatin domains and a smaller carboxyl-terminal domain. Fetuin-A is synthesized by the liver and secreted into blood stream, where its concentration in adult mammals ranges from 0.5 – 1.0 g/L. Fetuin-A occurs in high serum concentration during fetal life and involves in protease inhibitory activities and development-associated regulation of calcium metabolism and osteogenesis. It accumulates in bones and teeth as a major fraction of noncollagenous bone proteins Biologically, studies have demonstrated that Fetuin-A is the major calcification inhibitor found in circulation, where it interferes with calcium salt precipitation. Recent study has indicated that Fetuin-A level drops in uremic patients on hemodialysis in comparison to normal healthy controls. The low Fetuin-A level in patients with chronic kidney failure strongly associates with a higher cardiovascular mortality. On the other hand, it is demonstrated that a higher than normal serum Fetuin-A in older population associates incident diabetes, which is independent from other markers of insulin resistance. Further, a higher Fetuin-A level may be an independent risk marker of patients with for cardiovascular diseases.
This ELISA is designed, developed and produced for the quantitative measurement of human Fetuin-A in serum samples. The assay utilizes the two-site “sandwich” technique with two selected goat anti-human Fetuin-A polyclonal antibodies that bind to different epitopes of human Fetuin-A.
Assay standards, controls and prediluted patient serum samples containing human Fetuin-A is added to microtiter wells of microplate that was coated with a high affinity polyclonal goat anti-human Fetuin-A antibody. After the first incubation period, the antibody on the wall of microtiter well captures human Fetuin-A in the sample and unbound proteins in each microtiter well is washed away. Then a horseradish peroxidase （HRP） conjugated polyclonal anti-human Fetuin-A antibody is added to each microtiter well and a “sandwich” of “capture antibody - human Fetuin-A - HRP conjugated tracer antibody” is formed. The unbound tracer antibody is removed in the subsequent washing step. HRP conjugated tracer antibody bound to the well is then incubated with a substrate solution in a timed reaction and then measured in a spectrophotometric microplate reader. The enzymatic activity of the tracer antibody bound to the Fetuin-A on the wall of the microtiter well is directly proportional to the amount of Fetuin-A in the sample. A standard curve is generated by plotting the absorbance versus the respective human Fetuin-A concentration for each standard on point-to-point or cubical scales. The concentration of human Fetuin-A in test samples is determined directly from this standard curve.
REAGENTS: Preparation
1. Fetuin-A Antibody Coated Microplate ：96 wells
2. Fetuin-A Tracer Antibody ：1vial× 0.6 mL
3. Tracer Antibody Diluent：1vial× 12mL
4. Fetuin-A Assay Buffer Concentrate ：1vial× 11mL
5. ELISA Wash Concentrate，30X：1 bottle× 20mL
6. ELISA HRP Substrate ：1 bottle× 12mL
7. ELISA Stop Solution：1 bottle× 12mL
8. Fetuin-A Standards ：5 vials
9. Fetuin-A Controls ：2 vials
