Cat No.: 31060 Other Names: PRBP, RBP, PRO2222
Introduction
Retinol binding protein 4(RBP4), originally known as a specific transport of retinol in blood, is also a novel inflammatory and insulin
resistance marker. Serum RBP4 levels are elevated in insulin-resistant mice and humans with obesity and type 2 diabetes. Animal
experiments found that increased secretion of RBP4 might reduces insulin-dependent glucose uptake by muscle tissue by reducing
the activity of PI(3)K (phosphoinositide 3-kinase), and increased hepatic glucose output by increasing the expression of the enzyme PEPCK2 . Studies suggested that elevated serum RBP4 was associated with components of the metabolic syndrome, including
increased body-mass index, waist-to-hip ratio, serum triglyceride levels, and systolic blood pressure and decreased highdensity
lipoprotein cholesterol levels. Furthermore, circulating RBP4 concentrations were associated with subclinical cardiovascular disease, which imply that RBP4 could be involved in the development of atherosclerosis.
Principle of the Assay
This assay is a quantitative sandwich ELISA using monoclonal antibodies against human RBP4. The immunoplate is pre-coated with a monoclonal antibody specific for human RBP4 and the nonspecific binding sites are blocked. Standards and samples are pipetted
into the wells and any Human RBP4 present is bound by the immobilized antibody. After washing away any unbound substances, a
horseradish peroxidase (HRP)-linked monoclonal antibody specific for human RBP4 is added to the wells. After wash step to remove any unbound reagents, an HRP substrate solution is added and colour develops in proportion to the amount of human RBP4 bound
initially. The assay is stopped and the optical density of the wells determined using a microplate reader. Since the increases in
absorbance are directly proportional to the amount of captured human RBP4, the unknown sample concentration can be interpolated from a reference curve included in each assay.
Assay Performance
A. Typical representation of standard curve
The following standard curve is provided for demonstration only. A standard curve should be generated for each set of sample
assay.
RBP4 (ng/mL) | Absorbance (450 nm) | Blanked Absorbance |
0 | 0.091 | 0 |
3.12 | 0.146 | 0.055 |
6.25 | 0.194 | 0.103 |
12.5 | 0.30 | 0.209 |
25 | 0.504 | 0.413 |
50 | 0.836 | 0.745 |
100 | 1.586 | 1.495 |
200 | 2.499 | 2.408 |
B. Sensitivity
The lowest level of RBP4 that can be detected by this assay is 3.12 ng/mL.
C. Specificity
The antibody pair used in this assay is specific to human RBP4 and does not cross-react with mouse and rat RBP4, and other
cytokine or hormone molecules.
D. Precision
Intra-assay Precision (Precision within an assay)
Three samples of known concentration were tested 16 times on one plate.
Sample | Mean (μg/mL) | SD (μg/mL) | CV (%) |
1 | 18.03 | 0.654 | 3.63 |
2 | 49.07 | 1.158 | 2.36 |
3 | 3.91 | 0.117 | 2.99 |
Inter-assay Precision (Precision between assays)
Three samples of known concentration were tested in 10 separate assays.
Sample | Mean (μg/mL) | SD (μg/mL) | CV (%) |
1 | 6.53 | 0.284 | 4.35 |
2 | 17.11 | 0.524 | 3.06 |
3 | 44.21 | 1.154 | 2.61 |
E. Recovery
Serum samples were spiked with different amounts of human RBP4 and assayed.
Sample | Average % Recovery | Range % |
Serum (n=4) | 102 | 92-115 |
Publications Citing This Product
1. Alkharfy KM, Al-Daghri NM, Vanhoutte PM, Krishnaswamy S, Xu A. Serum retinol-binding protein 4 as a marker for cardiovascular disease in women. PloS one. 2012 Oct 31;7(10):e48612.
2. Ong KL, Rye KA, O'Connell R, Jenkins AJ, Brown C, Xu A, Sullivan DR, Barter PJ, Keech AC, FIELD Study Investigators. Long-
term fenofibrate therapy increases fibroblast growth factor 21 and retinol-binding protein 4 in subjects with type 2 diabetes. The
Journal of Clinical Endocrinology & Metabolism. 2012 Dec 1;97(12):4701-8.
3. Jüllig M, Yip S, Xu A, Smith G, Middleditch M, Booth M, Babor R, Beban G, Murphy R. Lower fetuin-A, retinol binding protein 4 and several metabolites after gastric bypass compared to sleeve gastrectomy in patients with type 2 diabetes. PLoS One. 2014 May 6;9(5):e96489.
4. Chen DL, Liess C, Poljak A, Xu A, Zhang J, Thoma C, Trenell M, Milner B, Jenkins AB, Chisholm DJ, Samocha-Bonet D.
Phenotypic characterization of insulin-resistant and insulin-sensitive obesity. The Journal of Clinical Endocrinology & Metabolism.
2015 Nov 1;100(11):4082-91.
5. Chen DL, Brown R, Liess C, Poljak A, Xu A, Zhang J, Trenell M, Jenkins A, Chisholm D, Samocha-Bonet D, Macefield VG. Muscle sympathetic nerve activity is associated with liver insulin sensitivity in obese non-diabetic men. Frontiers in physiology. 2017 Feb 28;8:101.
6. Wong YK, Cheung CY, Tang CS, Au KW, Hai JS, Lee CH, Lau KK, Cheung BM, Sham PC, Xu A, Lam KS. Age-biomarkers-clinical risk factors for prediction of cardiovascular events in patients with coronary artery disease. Arteriosclerosis, thrombosis, and vascular biology. 2018 Oct;38(10):2519-27.
7. Xiao Y, Xiao X, Xu A, Chen X, Tang W, Zhou Z. Circulating adipocyte fatty acid-binding protein levels predict the development of
subclinical atherosclerosis in type 2 diabetes. Journal of Diabetes and its Complications. 2018 Dec 1;32(12):1100-4.
8. Tang A, Coster AC, Tonks KT, Heilbronn LK, Pocock N, Purtell L, Govendir M, Blythe J, Zhang J, Xu A, Chisholm DJ. Longitudinal changes in insulin resistance in normal weight, overweight and obese individuals. Journal of clinical medicine. 2019 May;8(5):623.
9. Harari A, Coster AC, Jenkins A, Xu A, Greenfield JR, Harats D, Shaish A, Samocha-Bonet D. Obesity and insulin resistance are
inversely associated with serum and adipose tissue carotenoid concentrations in adults. The Journal of nutrition. 2020 Jan 1;150(1):
38-46.