AN ALTERNATIVE DIAGNOSTIC DESIGN FOR CHRONIC KIDNEY DISEASE DETECTION BASED ON CYSTATIN C
Objective: Chronic Kidney Disease (CKD) is usually diagnosed by measuring the glomerular filtration rate (GFR) and diagnostics are still inadequate at the clinical level. Most of the diagnostic kits available estimate GFR rate by determining clearance of serum creatinine using various instrumentation generally available at hospitals. Serum creatinine is considered the major marker for renal deficiency disorders. Additionally, it is also an indicator of muscle mass and dietary intake. Hence, the need for a more reliable marker for CKD arises. A low molecular weight protein cystatin C has been found to be a reliable biomarker for detection of kidney function as it is solely filtered by the glomerulus and not secreted by renal tubules.
Method: The basic set up of the kit was designed using a syringe containing multi walled carbon nanotube (MWCNT) conjugated protease. Casein beads were immersed inÂ PBS buffer in the syringe. The Glass/MWCNT/papain solid support was subsequently inserted into the syringe in such a way, that the beads came in contact with the immobilized enzyme conjugate. The inhibitory action of cystatin C against protease forms the basis for the functioning of the kit
Results: Results indicated that papain while immobilization needs to be in dynamic conformation. At 37 C gave better activity as compared to protein immobilized at 4C. FTIR observations confirmed the physical adsorption on the MWCNTs. The experimentation confirmed the feasibility of using prototype for detection of cystatin C.
Discussion: Papain conjugated with MWCNT indicated its temperature and pH stability. The initial design of the diagnostic kit for the detection of CKD has shown to be successful with a good detection range corresponding to stage I and II of CKD. Further testing needs to be done for the prototype using patient samples.
Keywords: Chronic kidney disease, Diagnostic kit, Immobilized papain, Protease inhibitor.Â
 Kellum JA, Levin N, Bouman C, Lameire N. Developing a consensus classification system for acute renal failure. Curr Opin Crit Care 2002;8:509â€“14. doi:10.1097/00075198-200212000-00005..
 WHO. Global status report on noncommunicable diseases. 2010. doi:ISBN 978 92 4 156422 9.
 Couser WG, Remuzzi G, Mendis S, Tonelli M. The contribution of chronic kidney disease to the global burden of major noncommunicable diseases. Kidney Int 2011;80:1258â€“70. doi:10.1038/ki.2011.368.
 Veerappan I, Abraham G. Chronic kidney disease: Current status, challenges and management in India. Ch. 130, Sec. 17:apiindia. org/medicine_update_2013. p. 593-7.
 WebMD.Chronic Kidney Disease; available from http://www.webmd.com/a-to-z-guides/chronic-kidney-disease-cause. WebMD Medical Reference from Healthwise 2013.
 Fassett RG, Venuthurupalli SK, Gobe GC, Coombes JS, Cooper MA, Hoy WE. Biomarkers in chronic kidney disease: a review. Kidney Int 2011;80:806â€“21. doi:10.1038/ki.2011.198.
 RANDOX renal function test available from http://www.randox.com/brochures/PDF%20 Brochure/ LT031. pdf. RANDOX.
 Mussap M, Plebani M. Biochemistry and clinical role of human cystatin C. Crit Rev Clin Lab Sci 2004; 41 467â€“550. doi:10.1080/10408360490504934.
 Hall YN, Choi AI, Chertow GM, Bindman AB. Chronic kidney disease in the urban poor. Clin J Am Soc Nephrol 2010;5:828â€“35. doi:10.2215/CJN.09011209.
 Qutb A, Syed G, Tamim HM, Al Jondeby M, Jaradat M, Tamimi W, Al Ghamdi G, Al Qurashi S, Flaiw A, Hejaili F, Al Sayyari AA. Cystatin C-based formula is superior to MDRD, Cockcroft-Gault and Nankivell formulae in estimating the glomerular filtration rate in renal allografts. Exp Clin Transplant.2009;7(4):197-202
 Petrovic S, Bogavac-Stanojevic N, Lakic D, Peco-Antic A, Vulicevic I, Ivanisevic I, Jelic-Ivanovic Z. Cost-effectiveness analysis of acute kidney injury biomarkers in pediatric cardiac surgery. Biochemia Medica. 2015; 25:262â€“271. doi:10.11613/BM.2015.027
 Wang Q, Zhou L, Jiang Y, Gao J. Improved stability of the carbon nanotubes-enzyme bioconjugates by biomimetic silicification. Enzyme Microb Technol 2011;49:11â€“6. doi:10.1016/j.enzmictec.2011.04.007.
 Hart SR, Waterfield MD, Burlingame AL, Cramer R. Factors governing the solubilization of phosphopeptides retained on ferric NTA IMAC beads and their analysis by MALDI TOFMS. J Am Soc Mass Spectrom 2002;13:1042â€“51. doi:10.1016/S1044-0305(02)00432-4.
 Ding L, Yao Z, Li T, Yue Q, Chai J. Study on Papain Immobilization on a Macroporous Polymer Carrier. Turkish J Chem 2003;27:627â€“37.
Sangeetha K, Abraham TE. Chemical modification of papain for use in alkaline medium. J Mol Catal B Enzym 2006;38:171â€“7. doi:10.1016/j.molcatb.2006.01.003.
 Zati Iwani AK , Ruziana Mona WZ, Nor Idayu R, Wan Nazaimoon WM. The Usefulness of Cystatin C as a Marker for Chronic Kidney Disease Universal Journal of Clinical Medicine 2013 1 (2): 28-33, DOI: 10.13189/ujcm.2013.010203
 Benaki DC, Anggeli A, Chryssikos GD, Yiannopoulos YD, Kamitsos EI, Brumley E, Case ST, Boden N, Hamodrakas SJ. Laser-raman and FT-IR spectroscopic studies of peptide analogues of silkmoth chorion protein segments. Int. J. Biol. Macromol. 1998; 23: 49â€“59.
 Nandy SK , Bhuyan R , Seal A. Modelling family 2 cystatins and their interaction with papain
Journal of Biomolecular Structure and Dynamics 2013; 31: 649-664
 Sangeetha K, Abraham TE. Chemical modification of papain for use in alkaline medium. J Mol Catal B Enzym 2006;38:171â€“7. doi:10.1016/j.molcatb.2006.01.003.
 Zati Iwani AK , Ruziana Mona WZ, Nor Idayu R, Wan Nazaimoon WM. The Usefulness of Cystatin C as a Marker for Chronic Kidney Disease Universal Journal of Clinical Medicine 2013 1(2): 28-33, DOI: 10.13189/ujcm.2013.010203
Moo Yong Park, Soo Jeong Choi, Jin Kuk Kim SDH andYong WL. Urinary cystatin C levels as a diagnostic and prognostic biomarker in patients with acute kidney injury. Nephrology 2013;18:256â€“62. doi:10.1111/nep.12037.
Li J, Dunn W, Breaud A, Elliott D, Sokoll LJ, Clarke W. Analytical performance of 4 automated assays for measurement of cystatin C. Clin Chem 2010;56:1336â€“9. doi:10.1373/clinchem.2009.141531.
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