Human Endothelin-1 QuantiGlo ELISA Kit

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QET00B
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Human Endothelin-1 Chemiluminescent ELISA Standard Curve
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Product Details
Citations (48)
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Human Endothelin-1 QuantiGlo ELISA Kit Summary

Assay Length
4.5 hours
Sample Type & Volume Required Per Well
Cell Culture Supernates (100 uL), Serum (100 uL), EDTA Plasma (100 uL), Urine (100 uL)
Sensitivity
0.102 pg/mL
Assay Range
0.3 - 250 pg/mL (Cell Culture Supernates, Serum, EDTA Plasma, Urine)
Specificity
Natural and synthetic human Endothelin-1
Cross-reactivity
Cross-reactivity observed with 1 or more available related molecules.Cross-species reactivity not tested.
Interference
No significant interference observed with available related molecules.

Product Summary

The QuantiGlo Human Endothelin-1 Chemiluminescent Immunoassay is a 4.5 hour solid phase ELISA designed to measure human Endothelin-1 levels in cell culture supernates, serum, EDTA plasma, and urine without extraction. It contains synthetic human Endothelin-1 and antibodies raised against the synthetic factor. This immunoassay has been shown to accurately quantitate human Endothelin-1. Results obtained using natural human Endothelin-1 showed dose curves that were parallel to the standard curves obtained using the QuantiGlo kit standards. These results indicate that this kit can be used to determine relative mass values of natural human Endothelin-1.

Recovery

The recovery of synthetic human Endothelin-1 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type Average % Recovery Range %
Cell Culture Media (n=4) 101 93-107
EDTA Plasma (n=4) 94 88-99
Serum (n=4) 95 87-103
Urine (n=4) 98 92-105

Linearity

To assess the linearity of the assay, samples containing or spiked with high concentrations of Endothelin-1 in various matrices were diluted with Calibrator Diluent to produce samples with values within the dynamic range of the assay.
Human Endothelin-1 Chemiluminescent ELISA Linearity

Scientific Data

Human Endothelin-1 Chemiluminescent ELISA Standard Curve

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Preparation and Storage

Shipping
The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.

Background: Endothelin-1

Endothelin-1 (ET-1), a peptide of 21 amino acid (aa) residues, is a pleiotropic molecule best known for its action as a potent vasoconstrictor (1). Originally isolated from porcine aortic endothelial cells, ET-1 is one of a family of three proteins encoded by distinct genes that also includes Endothelin-2 (ET-2) and Endothelin-3 (ET-3) (2, 3). ET-2 and ET-3 differ from ET-1 by 2 and 6 amino acids, respectively (1, 2). All members of the Endothelin family contain two essential disulfide bridges and six conserved aa residues at the C-terminus. Human ET-1 is initially synthesized as a pre-pro-polypeptide of 212 amino acids (2, 4). It is proteolytically cleaved by a signal peptidase to produce pro-ET-1 and further processed by a Furin-like protease to yield a 38 aa peptide termed Big ET-1 (5, 6). Big ET-1 is then cleaved by the membrane-bound metalloprotease Endothelin-converting enzyme (ECE-1), producing the potent 21 aa mature form ET-1 (aa 1-21) (7, 8). Alternatively, ET-1 may exist in an active 31 aa form (ET-1 (aa 1-31)) following cleavage of Big ET-1 by chymase (9-12). The vascular endothelium is an abundant source of ET-1 (3, 13). It may also be expressed by leukocytes, smooth muscle cells, mesangial cells, cardiac myocytes, and astrocytes (14, 15). ET-1 can be induced in endothelial cells by many factors including mechanical stimulation, various hormones, and pro-inflammatory cytokines (16). Production is inhibited by nitric oxide (NO), Prostacyclin, and atrial natriuretic peptide (ANP) (17-19). 

Two receptors for the Endothelin family have been cloned and designated ETA and ETB (20-23). ETA and ETB belong to the large family of heptahelical G protein-coupled receptors. The ETA receptor shows a higher affinity for ET-1 than for ET-2 and lowest affinity for ET-3, while the ETB receptor shows approximately equal affinity for each of the three Endothelins (21, 22, 24). ETA is primarily responsible for the vasoconstrictor effects of ET-1 and is expressed by blood vessel smooth muscle cells (25, 26). The ETB receptor is also present in smooth muscle and the endothelia of blood vessels, kidney, lung, and brain (27). ET-1 has the ability to activate an array of signaling cascades including classical phosphatidylinositol turnover pathways leading to downstream PKC activation and Ca2+ mobilization (28-32). Other potential signaling mediators activated or produced by ET-1 include PI 3-kinase/Akt, NO, FAK, and Rho GTPases (32-37). ET-1 signaling may also be mediated indirectly via transactivation of the EGF receptor leading to downstream signaling by Ras and MAP kinases (38, 39). Injection of a single dose of ET-1 produces an initial decrease in systemic blood pressure followed by a prolonged increase in blood pressure (16, 40, 41). Blockade of Endothelin receptors with a systemic injection of an ETA/ETB antagonist causes progressive vasodilation, and elevated levels of ET-1 are found in some forms of human hypertension (42, 43). ET-1 also stimulates cardiac contraction and the growth of cardiac myocytes, regulates the release of vasoactive substances, and stimulates smooth muscle cell mitogenesis (32, 44-46). It also acts as a pro-survival factor for endothelial cells and regulates secretion by hypothalamic and pituitary cells (47, 48). ET-1 may control inflammatory responses by promoting the adhesion and migration of neutrophils and stimulating the production of pro-inflammatory cytokines (49-53). It has also been implicated in cancer progression at several levels including regulating the proliferation and migration of tumor cells and acting as a pro-angiogenic factor (54, 55). In addition, ET-1 has putative roles in other pathologies including septic shock, atherosclerosis, heart failure, renal insufficiency, pulmonary hypertension, and cerebrovascular conditions associated with subarachnoid hemorrhage (15, 56-63).

Entrez Gene IDs:
1906 (Human); 13614 (Mouse); 24323 (Rat)
Alternate Names:
ARCND3; EDN1; endothelin 1; Endothelin-1; ET1; HDLCQ7; PPET1; preproendothelin-1; QME

Citations for Human Endothelin-1 QuantiGlo ELISA Kit

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

48 Citations: Showing 1 - 10
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  1. Transpulmonary Plasma Endothelin-1 Arterial:Venous Ratio Differentiates Survivors from Non-Survivors in Critically Ill Patients with COVID-19-Induced Acute Respiratory Distress Syndrome
    Authors: Vassiliou, AG;Roumpaki, A;Keskinidou, C;Athanasiou, N;Tsipilis, S;Jahaj, E;Vrettou, CS;Giannopoulou, V;Halioti, A;Ferentinos, G;Dimopoulou, I;Kotanidou, A;Langleben, D;Orfanos, SE;
    International journal of molecular sciences
    Species: Human
    Sample Types: Plasma
  2. Effect of endothelin-1 on the blood pressure response to acute hypoxia and hyperoxia in healthy young men
    Authors: Gonsalves, AM;Baker, SE;Jacob, DW;Harper, JL;Manrique-Acevedo, CM;Limberg, JK;
    Physiological reports
    Species: Human
    Sample Types: Plasma
  3. Crucial Interactions between Altered Plasma Trace Elements and Fatty Acids Unbalance Ratio to Management of Systemic Arterial Hypertension in Diabetic Patients: Focus on Endothelial Dysfunction
    Authors: Gouaref, I;Otmane, A;Makrelouf, M;Abderrhmane, SA;Haddam, AEM;Koceir, EA;
    International journal of molecular sciences
    Species: Human
    Sample Types: Plasma
  4. Myocardial transcriptomic analysis of diabetic patients with aortic stenosis: key role for mitochondrial calcium signaling
    Authors: Cherpaz, M;Meugnier, E;Seillier, G;Pozzi, M;Pierrard, R;Leboube, S;Farhat, F;Vola, M;Obadia, JF;Amaz, C;Chalabreysse, L;May, C;Chanon, S;Brun, C;Givre, L;Bidaux, G;Mewton, N;Derumeaux, G;Bergerot, C;Paillard, M;Thibault, H;
    Cardiovascular diabetology
    Species: Human
    Sample Types: Serum
  5. The Regulation of Endothelin-1 in Pregnancies Complicated by Gestational Diabetes: Uncovering the Vascular Effects of Insulin
    Authors: Fato, BR;Beard, S;Binder, NK;Pritchard, N;Kaitu'u-Lino, TJ;de Alwis, N;Hannan, NJ;
    Biomedicines
    Species: Human
    Sample Types: Plasma
  6. Interplay of Race and Neighborhood Deprivation on Ambulatory Blood Pressure in Young Adults
    Authors: Jeong, S;Linder, BA;Barnett, AM;Tharpe, MA;Hutchison, ZJ;Culver, MN;Sanchez, SO;Nichols, OI;Grosicki, GJ;Bunsawat, K;Nasci, VL;Gohar, EY;Fuller-Rowell, TE;Robinson, AT;
    medRxiv : the preprint server for health sciences
    Species: Human
    Sample Types: Urine
  7. Vascular Function, Systemic Inflammation, and Coagulation Activation 18 Months after COVID-19 Infection: An Observational Cohort Study
    Authors: LH Willems, LMC Jacobs, LA Groh, H Ten Cate, HMH Spronk, B Wilson-Sto, G Hannink, SMJ van Kuijk, C Ghossein-D, M Nagy, DHJ Thijssen, AS van Peters, MC Warlé
    Journal of Clinical Medicine, 2023-02-10;12(4):.
    Species: Human
    Sample Types: Plasma
  8. Dietary sodium restriction prevents vascular endothelial growth factor inhibitor-induced hypertension
    Authors: L van Doorn, WJ Visser, DCH van Dorst, KM Mirabito C, SLW Koolen, AVE de Mik, IM Garrelds, DM Bovée, EO de Hoop, S Bins, FALM Eskens, EJ Hoorn, AH Jan Danser, RHJ Mathijssen, J Versmissen
    British Journal of Cancer, 2022-11-10;0(0):1-9.
    Species: Human
    Sample Types: Plasma
  9. Endothelial dysfunction and altered endothelial biomarkers in patients with post-COVID-19 syndrome and chronic fatigue syndrome (ME/CFS)
    Authors: M Haffke, H Freitag, G Rudolf, M Seifert, W Doehner, N Scherbakov, L Hanitsch, K Wittke, S Bauer, F Konietschk, F Paul, J Bellmann-S, C Kedor, C Scheibenbo, F Sotzny
    Journal of Translational Medicine, 2022-03-22;20(1):138.
    Species: Human
    Sample Types: Serum
  10. Association of Plasma Endothelin-1 with Blood Pressure Progression Among African Americans: The Jackson Heart Study
    Authors: AD Kaze, X Gao, SK Musani, A Bidulescu, AG Bertoni, M Abdalla, JB Echouffo-T
    American heart journal, 2022-01-02;0(0):.
    Species: Human
    Sample Types: Serum
  11. Serum Metabolomic Analysis of Coronary Heart Disease Patients with Stable Angina Pectoris Subtyped by Traditional Chinese Medicine Diagnostics Reveals Biomarkers Relevant to Personalized Treatments
    Authors: N Guo, P Wang, J Yang, X Yang, M van der Vo, M Wildwater, J Wei, X Tang, M Wang, H Yang
    Frontiers in Pharmacology, 2021-06-14;12(0):664320.
    Species: Human
    Sample Types: Serum
  12. Adaptive response triggered by the repeated SCUBA diving is reflected in cardiovascular, muscular, and immune biomarkers
    Authors: M Žarak, A Perovi?, M Njire Brat, S Šupraha Go, J Dumi?
    Physiological Reports, 2021-01-01;9(2):e14691.
    Species: Human
    Sample Types: Plasma
  13. EDN1-AS, A Novel Long Non-coding RNA Regulating Endothelin-1 in Human Proximal Tubule Cells
    Authors: LG Douma, K Solocinski, SH Masten, DH Barral, SJ Barilovits, LA Jeffers, KD Alder, R Patel, CS Wingo, KD Brown, BD Cain, ML Gumz
    Front Physiol, 2020-03-13;11(0):209.
    Species: Human
    Sample Types: Cell culture supernate
  14. FURIN Expression in Vascular Endothelial Cells Is Modulated by a Coronary Artery Disease-Associated Genetic Variant and Influences Monocyte Transendothelial Migration
    Authors: X Yang, W Yang, DG McVey, G Zhao, J Hu, RN Poston, M Ren, K Willeit, S Coassin, J Willeit, TR Webb, NJ Samani, M Mayr, S Kiechl, S Ye
    J Am Heart Assoc, 2020-02-11;9(4):e014333.
    Species: Human
    Sample Types: Cell Culture Supernates
  15. Clinical Significance of High-Mobility Group Box 1 Protein (HMGB1) and Nod-Like Receptor Protein 3 (NLRP3) in Patients with Ulcerative Colitis
    Authors: Y Chen, D Wu, L Sun
    Med. Sci. Monit., 2020-01-05;26(0):e919530.
    Species: Human
    Sample Types: Serum
  16. Plasma endothelin-1 levels are increased in atrial fibrillation patients with hyperthyroidism
    Authors: F Mayyas, N Saadeh, K Al-Muqbel, DR Van Wagone
    PLoS ONE, 2018-12-04;13(12):e0208206.
    Species: Human
    Sample Types: Plasma
  17. DKK1 and Kremen Expression Predicts the Osteoblastic Response to Bone Metastasis
    Authors: KL Clines, GA Clines
    Transl Oncol, 2018-05-15;11(4):873-882.
    Species: Human
    Sample Types: Cell Culture Supernates
  18. Intervention of integrative medicine treatment has impact on serum levels of ET-1, TNF-?, MLT in RA-CVD
    Authors: M Chen, Z Li, Z Zhang, Y Du, Y Zhang, M Xu, C Sun
    Saudi J Biol Sci, 2018-05-14;25(5):959-964.
    Species: Human
    Sample Types: Serum
  19. Acute Pressor Response to Psychosocial Stress Is Dependent on Endothelium-Derived Endothelin-1
    Authors: BM Fox, BK Becker, AS Loria, KA Hyndman, C Jin, H Clark, R Johns, M Yanagisawa, DM Pollock, JS Pollock
    J Am Heart Assoc, 2018-02-16;7(4):.
    Species: Mouse
    Sample Types: Plasma
  20. Impaired endothelial function in patients with cryptogenic stroke and patent foramen ovale is not affected by closure
    Authors: M Lantz, K Kostulas, M Settergren, C Sjöstrand
    J Interv Cardiol, 2017-04-25;0(0):.
    Species: Human
    Sample Types: Serum
  21. Endothelin receptor-specific control of endoplasmic reticulum stress and apoptosis in the kidney
    Authors: C De Miguel, WC Hamrick, JL Hobbs, DM Pollock, PK Carmines, JS Pollock
    Sci Rep, 2017-02-23;7(0):43152.
    Species: Rat
    Sample Types: Plasma
  22. Free radical scavenging decreases endothelin-1 excretion and glomerular albumin permeability during type 1 diabetes
    Physiol Rep, 2016-12-01;4(24):.
    Species: Rat
    Sample Types: Urine
  23. Allergic Inflammation Leads to Neuropathic Pain via Glial Cell Activation
    Authors: Jun-Ichi Kira
    J. Neurosci., 2016-11-23;36(47):11929-11945.
    Species: Human
    Sample Types: Serum
  24. Effect of OSA on hypoxic and inflammatory markers during CPAP withdrawal: Further evidence from three randomized control trials
    Authors: Malcolm Kohler
    Respirology, 2016-11-18;0(0):.
    Species: Human
    Sample Types: Plasma
  25. Albuminuria Is Associated with Endothelial Dysfunction and Elevated Plasma Endothelin-1 in Sickle Cell Anemia
    PLoS ONE, 2016-09-26;11(9):e0162652.
    Species: Human
    Sample Types: Plasma
  26. p66Shc regulates renal vascular tone in hypertension-induced nephropathy
    Authors: Bradley Miller
    J Clin Invest, 2016-06-06;0(0):.
    Species: Rat
    Sample Types: Tissue Culture Supernates
  27. Endothelin-1 Predicts Hemodynamically Assessed Pulmonary Arterial Hypertension in HIV Infection.
    Authors: Parikh R, Ma Y, Scherzer R, Heringer A, Macgregor J, Martin J, Deeks S, Ganz P, Hsue P
    PLoS ONE, 2016-01-11;11(1):e0146355.
    Species: Human
    Sample Types: Plasma
  28. Association of endothelin-1 and matrix metallopeptidase-9 with metabolic syndrome in middle-aged and older adults.
    Authors: Yu A, Tam B, Yau W, Chan K, Yu S, Chung T, Siu P
    Diabetol Metab Syndr, 2015-12-10;7(0):111.
    Species: Human
    Sample Types: Serum
  29. Optimum AT1 receptor-neprilysin inhibition has superior cardioprotective effects compared with AT1 receptor blockade alone in hypertensive rats.
    Authors: Roksnoer L, van Veghel R, de Vries R, Garrelds I, Bhaggoe U, Friesema E, Leijten F, Poglitsch M, Domenig O, Clahsen-van Groningen M, Hoorn E, Jan Danser A, Batenburg W
    Kidney Int, 2015-04-01;88(1):109-20.
    Species: Rat
    Sample Types: Plasma
  30. Chemotherapy-induced pulmonary hypertension: role of alkylating agents.
    Authors: Ranchoux B, Gunther S, Quarck R, Chaumais M, Dorfmuller P, Antigny F, Dumas S, Raymond N, Lau E, Savale L, Jais X, Sitbon O, Simonneau G, Stenmark K, Cohen-Kaminsky S, Humbert M, Montani D, Perros F
    Am J Pathol, 2014-12-11;185(2):356-71.
    Species: Rabbit
    Sample Types: Plasma
  31. The in vitro pharmacology of GS-5759, a novel bifunctional phosphodiesterase 4 inhibitor and long acting beta2-adrenoceptor agonist.
    Authors: Tannheimer S, Sorensen E, Cui Z, Kim M, Patel L, Baker W, Phillips G, Wright C, Salmon M
    J Pharmacol Exp Ther, 2014-02-10;349(1):85-93.
    Species: Human
    Sample Types: Cell Culture Supernates
  32. Aberrant chloride intracellular channel 4 expression contributes to endothelial dysfunction in pulmonary arterial hypertension.
    Authors: Wojciak-Stothard B, Abdul-Salam V, Lao K, Tsang H, Irwin D, Lisk C, Loomis Z, Stenmark K, Edwards J, Yuspa S, Howard L, Edwards R, Rhodes C, Gibbs J, Wharton J, Zhao L, Wilkins M
    Circulation, 2014-02-06;129(17):1770-80.
    Species: Human
    Sample Types: Cell Culture Supernates
  33. miR-218 regulates focal adhesion kinase-dependent TGFbeta signaling in fibroblasts.
    Authors: Guo F, Carter D, Leask A
    Mol Biol Cell, 2014-02-05;25(7):1151-8.
    Species: Human
    Sample Types: Cell Culture Supernates
  34. Gingival fibroblasts display reduced adhesion and spreading on extracellular matrix: a possible basis for scarless tissue repair?
    Authors: Guo F, Carter DE, Mukhopadhyay A, Leask A
    PLoS ONE, 2011-11-02;6(11):e27097.
    Species: Human
    Sample Types: Cell Culture Supernates
  35. Daily red wine consumption improves vascular function by a soluble guanylyl cyclase-dependent pathway.
    Authors: Botden IP, Langendonk JG, Meima ME
    Am. J. Hypertens., 2010-11-18;24(2):162-8.
    Species: Human
    Sample Types: Plasma
  36. Early life stress downregulates endothelin receptor expression and enhances acute stress-mediated blood pressure responses in adult rats.
    Authors: Loria AS, D'Angelo G, Pollock DM, Pollock JS
    Am. J. Physiol. Regul. Integr. Comp. Physiol., 2010-04-21;299(1):R185-91.
    Species: Rat
    Sample Types: Plasma
  37. Endothelin receptors blockade blunts hypoxia-induced increase in PAP in humans.
    Authors: Pham I, Wuerzner G, Richalet JP, Peyrard S, Azizi M
    Eur. J. Clin. Invest., 2010-03-01;40(3):195-202.
    Species: Human
    Sample Types: Plasma
  38. Vehicular emissions induce vascular MMP-9 expression and activity associated with endothelin-1-mediated pathways.
    Authors: Lund AK, Lucero J, Lucas S, Madden MC, McDonald JD, Seagrave JC, Knuckles TL, Campen MJ
    Arterioscler. Thromb. Vasc. Biol., 2009-01-15;29(4):511-7.
    Species: Human
    Sample Types: Plasma
  39. Effects of nonoral estradiol-micronized progesterone or low-dose oral estradiol-drospirenone therapy on metabolic variables and markers of endothelial function in early postmenopause.
    Authors: Casanova G, Radavelli S, Lhullier F, Spritzer PM
    Fertil. Steril., 2008-08-15;92(2):605-12.
    Species: Human
    Sample Types: Plasma
  40. Endothelial function, endothelin-1, and fibrinogen in young women using the vaginal contraceptive ring.
    Authors: Torgrimson BN, Meendering JR, Miller NP, Kaplan PF, Minson CT
    Fertil. Steril., 2008-08-15;92(2):441-7.
    Species: Human
    Sample Types: Serum
  41. Regulation of angiogenic factors by HDM2 in renal cell carcinoma.
    Authors: Carroll VA, Ashcroft M
    Cancer Res., 2008-01-15;68(2):545-52.
    Species: Human
    Sample Types: Cell Culture Supernates
  42. Eosinophil-derived cationic proteins activate the synthesis of remodeling factors by airway epithelial cells.
    Authors: Pegorier S, Wagner LA, Gleich GJ, Pretolani M
    J. Immunol., 2006-10-01;177(7):4861-9.
    Species: Human
    Sample Types: Cell Culture Supernates
  43. Digital blood flow and plasma endothelin concentration in clinically endotoxemic horses.
    Authors: Menzies-Gow NJ, Bailey SR, Stevens K, Katz L, Elliott J, Marr CM
    Am. J. Vet. Res., 2005-04-01;66(4):630-6.
    Species: Equine
    Sample Types: Plasma
  44. Mechanisms of elevated plasma endothelin-1 in CHF: congestion increases pulmonary synthesis and secretion of endothelin-1.
    Authors: von Lueder TG, Kjekshus H, Edvardsen T, ØIe E, Urheim S, Vinge LE, Ahmed MS, Smiseth OA, Attramadal H
    Cardiovasc. Res., 2004-07-01;63(1):41-50.
    Species: Porcine
    Sample Types: Plasma
  45. Novel endothelin receptor antagonist attenuates endotoxin-induced lung injury in sheep.
    Authors: Kuklin VN, Kirov MY, Evgenov OV, Sovershaev MA, Sjoberg J, Kirova SS, Bjertnaes LJ
    Crit. Care Med., 2004-03-01;32(3):766-73.
    Species: Ovine
    Sample Types: Plasma
  46. Renal endothelin in chronic angiotensin II hypertension.
    Authors: Sasser JM, Pollock JS, Pollock DM
    Am. J. Physiol. Regul. Integr. Comp. Physiol., 2002-07-01;283(1):R243-8.
    Species: Rat
    Sample Types: Tissue Homogenates
  47. Passive transfer of scrub typhus plasma to patients with AIDS: a descriptive clinical study.
    Authors: Watt G, Kantipong P, Jongsakul K, de Souza M, Burnouf T
    QJM, 2001-11-01;94(11):599-607.
    Species: Human
    Sample Types: Plasma
  48. Endothelin and ET(A) receptors in long-term arterial pressure homeostasis in conscious nonhuman primates.
    Authors: Reinhart GA, 2018, Preusser LC, Opgenorth TJ, Wegner CD, Cox BF
    1364165, 2000-11-01;279(5):R1701-6.
    Species: Primate - Macaca fascicularis (Crab-eating Monkey or Cynomolgus Macaque)
    Sample Types: Plasma

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Average Rating: 4.7 (Based on 3 Reviews)

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Human Endothelin-1 QuantiGlo ELISA Kit
By Anonymous on 05/24/2019
Sample Tested: Plasma

Human Endothelin-1 QuantiGlo ELISA Kit
By Anonymous on 07/24/2017
Sample Tested: Serum

undiluted human serum from healthy donors


Human Endothelin-1 QuantiGlo ELISA Kit
By Anonymous on 06/29/2017
Sample Tested: EDTA Plasma

Very robust and easy to use kit.