Lipocalin 2/ NGAL Human ELISA

Summary

Lipocalin-2 (LCN2) is a 25 kDa secretory glycoprotein, also called NGAL (neutrophil gelatinase-associated lipocalin); NL (neutrophil lipocalin); p25; oncogen 24p3 and 25 kDa alpha-2-microglobulin-related subunit of MMP-9 (LCN2 forms a covalently linked, disulfide-bridged heterodimer with the 92 kDa type V collagenase (MMP-9)). LCN2 is predominantly expressed in adipose tissue and liver. It belongs to the lipocalin superfamily that consists of over 20 small secretory proteins. Lipocalin folds consist of 8 antiparallel ß-sheets that surround a hydrophobic pocket. A common feature of this protein family, following from its structure, is its capacity to bind and transport small lipophilic substancies such as free fatty acids, retinoids, arachidonic acid and various steroids. Although Lipocalin-2 was identified more than a decade ago, the physiologic function of this protein remains poorly understood. LCN2 appears to be upregulated in cells under the “stress” (e.g. from infection, inflammation, in tissues undergoing involution to ischemia or neoplastic transformation). Plasma levels of LCN2 rise in inflammatory or infective condition. It mediates an immune response to bacterial infection by sequestering iron. In this case, LCN2 may represent a promising candidate as a therapeutic agent against bacterial infection. Several recent reports suggest that LCN2 might represent a sensitive biomarker for early renal injury. In cardiopulmonary bypass-induced acute renal injury and cisplatin-induced nephrotoxic injury, increased de novo synthesis of LCN2 in proximal tubule cells leads to sharply increased concentration of this protein in both urine and serum. LCN2 might also be critical for normal kidney formation in the earliest stages of mammalian development. LCN2 may play an important role in breast cancer, in complex with MMP-9, by protecting MMP-9 from degradation thereby enhancing its enzymatic activity and facilitating angiogenesis and tumor growth. LCN2 is also highly expressed after malignant transformation of the lung, colon and pancreatic epithelia. Circulating levels of LCN2 play a causative role in pathogenesis of obesity-induced metabolic disorders such as insulin resistance, Type 2 Diabetes Mellitus and cardiovascular disorders. In addition, serum LCN2 concentrations were positively associated with adipocyte-fatty acid binding protein (A-FABP), a novel serum marker for adiposity and metabolic syndrome.

Areas of investigation: Bacterial infection Renal injury Angiogenesis Oncology Diabetes mellitus Metabolic syndrome

Features

• It is intended for research use only
• The total assay time is less than 3.5 hours.
• The kit measures Lipocalin-2 in serum, plasma (EDTA, citrate, heparin) and urine.
• Assay format – 96 wells
• Quality controls are human serum based. No animal sera are used.
• Standard is recombinant protein based.
• Components of kit are provided ready to use, concentrated or lyophilized.

Research topic

Energy metabolism and body weight regulation, Renal disease

Assay format

Sandwich ELISA, Biotin-labelled antibody

Applications

Plasma-Citrate, Plasma-EDTA, Plasma-Heparin, Serum, Urine

Sample requirements

10 µl/well

Storage/Shipping

Store the kit at 2–8°C. Under these conditions, the kit is stable until the expiration date (see label on the box).

Calibration Curve

0.3 – 10 ng/ml

Calibration range

0.3 to 10 ng/ml

Limit of detection

Limit of Detection (LOD) (defined as concentration of analyte giving absorbance higher than mean absorbance of blank* plus three standard deviations of the absorbance of blank: Ablank + 3×SDblank) is calculated from the real Lipocalin-2 values in wells and is 0.02 ng/ml

Intra-assay (Within-Run, n=8)

CV = 7.71 %

Inter-assay (Run-to-Run, n=7)

CV = 9.75 %

Spiking Recovery

98.0 %

Dilution Linearity

104.5 %

Cross-Reactivity

Monkey

References to summary

• Bachorzewska-Gajewska H., Malyszko J., Sitniewska E., Malyszko J.S., Poniatowski B., Pawlak K and Dobrzycki S.: NGAL (neutrophil gelatinase-associated lipocalin) and cystatin C: Are they good predictors of contrast nephropathy after percutaneous coronary interventions in patients with stable angina and normal serum creatinine? Int J Cardiol. 2007
• Bachorzewska-Gajewska H., Malyszko J., Sitniewska E., Malyszko J.S., Dobrzycki S.: Neutrophil gelatinase-associated lipocalin (NGAL) correlations with cystatin C, serum creatinine and eGFR in patients with normal serum creatinine undergoing coronary angiography. Nephrol Dial Transplant. 2007 Jan;22(1):295–6
• Lin H.H., Li W.W., Lee Y.C., Chu S.T.: Apoptosis induced by uterine 24p3 protein in endometrial carcinoma cell line. Toxicology 2007; 234 (3):203–15
• Mitsnefes M.M, Kathman T.S., Mishra J., Kartal J., Khoury P.R., Nickolas T.L., Barasch J., Devarajan P.: Serum neutrophil gelatinase-associated lipocalin as a marker of renal function in children with chronic kidney disease. Pediatr Nephrol. 2007;22(1):101–8
• Mori K and Nakao K.: Neutrophil gelatinase-associated lipocalin as the real-time indicator of active kidney damage. Kidney Int. 2007;71(10):967–70
• Nguyen M.T., Devarajan P.: Biomarkers for the early detection of acute kidney injury. Pediatr Nephrol. 2007
• van Dam R.M and Hu F.B.: Lipocalins and Insulin Resistance: Etiological Role of Retinol-Binding Protein 4 and Lipocalin-2. ClinChem 2007; 53:1
• Berger T, Togawa A.,Duncan G.S., Elia A.J.,You-Ten A., Wakeham A., Fong H.E.H., Cheung C.C. and Mak T.W.: Lipocalin-2-deficient mice exhibit increased sensitivity to Escherichia colli infection but not to ischemia-reperfusion injury. Proc Natl Acad Sci U S A 2006;10­3(6):1834–9
• Brunner H.I., Mueller M., Rutherford C., Passo M.H., Witte D., Grom A., Mistra J. and Devarajan P.: Urinary neutrophil gelatinase-associated lipocalin as a biomarker of nephritis in childhood-onset systemic lupus erythematosus. Arthritis Rheum 2006;54(8):2577–84
• Mishra J., Ma Q., Kelly C., Mitsnefes M., Mori K., Barasch J., Devarajan P.: Kidney NGAL is a novel early marker of acute injury following transplantation. Pediatr Nephrol. 2006;21(6):856–63
• Parikh C. R., Jani A., Mishra J., Ma Q., Kelly C., Barasch J., Edelstein C. L. and Devarajan P.: Urine NGAL and IL-18 are predictive biomarkers for delayed graft function following kidney transplantation. Am J Transplant. 2006;6(7):1639–45
• Trachtman H., Christen E., Cnaan A., Patrick J., Mai V., Mistra J., Jain A., Bullington N., Devarajan P.: Urinary neutrophil gelatinase-associated lipocalcin in D+HUS: a novel marker of renal injury. Pediatr Nephrol. 2006;21(7):989–94
• Vaidya V.S., Ramirez V., Ichimura T., Bobadilla N.A. and Bonventre J.V.: Urinary kidney injury molecule-1: a sensitive quantitative biomarker for early detection of kidney tubular injury. Am J Physiol Renal Physiol. 2006;290(2):F517–29
• Wang Y., Lam K.S.L., Kraegen E.W., Sweenery G., Zhang J., Tso A.W.K., Chow W-S., Wat N.M.S., Xu J.Y., Hoo R.C.L and Xu A. : Lipocalin-2 an inflammatory Marker Closely Associated with Obesity, Insulin Resistance and Hyperglycemia in Humans. ClinChem 2006 53(1):34–41
• Miharada K., Hiroyama T., Sudo K., Nagasawa T. and Nakamura Y.: Lipocalin-2 function as a negative regulator of red blood cell production in an autocrine fasion. FASEB Journal 2005:19; 1881–3
• Flo T.H., Smith K.D., Rodriguez D.J., Holmes M.A., Strong R.K. at al.: Lipocalin-2 mediates an immune response to bacterial infection by sequestrating iron. Nature 2004;432(7019):917–21
• Mishra J., Ma Q., Prada A., Mitsnefes M., Zahedi K., Yang J., Barasch J. and Devarajan P.: Identification of neutrophil gelatinase-associated lipocalin as a novel early urinary biomarker for ischemic renal injury. J Am Soc Nephrol. 2003 (10):2534–43

By research topic:

• Energy metabolism and body weight regulation
• Renal disease

By molecule:

• Lipocalin-2/NGAL

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