Targeted Mass Spectrometric Immunoassay for Detection of Cystatin C Isoforms in Cerebrospinal Fluid

Nedelkov, Dobrin; Shaik, Shainag; Trenchevska, Olgica; Aleksovski, Vasko; Mitrevski, Angel; Стојаноски, Киро

doi:10.2174/1875039700801010054

Cited by 7 publications

(8 citation statements)

References 12 publications

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“…In addition to CKD, CysC has been related to CVD and different neurological disorders. While higher levels of circulating CysC are associated with increased CVD risk, , increased expression and secretion in the brain may have both neurodegenerative as well as neuroprotective effects. , Proteoforms of CysC have been identified in blood samples and cerebrospinal fluids ,, and may give further insights into pathologies of CKD , and neurological disorders.…”

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confidence: 99%

Multiplex Immuno-MALDI-TOF MS for Targeted Quantification of Protein Biomarkers and Their Proteoforms Related to Inflammation and Renal Dysfunction

et al. 2018

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Circulating proteins are widely used as biomarkers in clinical applications for the diagnosis, prediction, and treatment of numerous diseases. Immunoassays are the most common technologies for quantification of protein biomarkers and exist in various formats. Traditional immunoassays offer sensitive and fast analyses but cannot differentiate between proteoforms. Protein microheterogeneity, mainly due to post-translational modification, has been recognized as a fingerprint for different pathologies, and knowledge about proteoforms is an important step toward personalized medicine. Mass spectrometry (MS) has emerged to be a powerful technique for the characterization and quantification of proteoforms. We have established a novel four-plex immunoassay based on Matrix-Assisted Laser Desorption/Ionization Time-Of-Flight (MALDI-TOF) MS for the targeted quantification of the inflammatory markers C-reactive protein (CRP), serum amyloid A (SAA), and calprotectin (S100A8/9) and the kidney function marker cystatin C (CysC). Antibodies were covalently bound to superparamagnetic beads, which delivered robust and fast sample processing. Polyhistidine-tagged recombinant target proteins were used as internal standards for quantification. Our method identified a number of proteoforms for SAA ( n = 11), S100A8/9 ( n = 4) and CysC ( n = 4). The assay was characterized by low limits of detection (0.01-0.06 μg/mL) and low coefficients of variation (3.8-9.4%). Method validation demonstrated good between-assay agreement with immuno-turbidimetry ( R = 0.963 for CRP), ELISA ( R = 0.958 for SAA; R = 0.913 for S100A8/9), and nephelometry ( R = 0.963 for CysC). The low sample consumption of 20 μL and the high sample throughput of 384 samples per day make this targeted immuno-MALDI approach suited for assessment of inflammatory and renal status in large cohort studies based on precious biobanks samples.

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confidence: 99%

Multiplex Immuno-MALDI-TOF MS for Targeted Quantification of Protein Biomarkers and Their Proteoforms Related to Inflammation and Renal Dysfunction

et al. 2018

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“…Moreover, paired serum/CSF samples origin from same individuals was screened. As presented in the previous studies, using the optimized MSIA qualitative assay, cysC and its wild type isoforms were detected in the analyzed CSF samples (Nedelkov, Shaik et al 2008). However, several correlations were observed when matched serum/CSF samples were analyzed.…”

Section: Qualitative Cysc Msia Assay In Csf and Serummentioning

confidence: 58%

“…8. MALDI-TOF MS spectra obtained after cystatin C MSIA in a) serum sample (control); b) CSF sample (control); c) paired serum/CSF sample (control) and d) paired serum/CSF sample (patients with diagnosed multiple sclerosis) (reprinted with permission from (Nedelkov, Shaik et al 2008))…”

Section: Qualitative Cysc Msia Assay In Csf and Serummentioning

confidence: 99%

Developing Novel Methods for Protein Analysis and Their Potential Implementation in Diagnosing Neurological Diseases

Trenchevska¹,

Aleksovski²,

Nedelkov³

et al. 2012

Advanced Topics in Neurological Disorders

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“…Using immunoaffinity pipettes it was possible to extract only the targeted protein and to analyze it with MALDI-TOF-MS. Experimental conditions such as sample dilution and incubation time were optimized in order to provide better results (9).…”

Section: Methodsmentioning

confidence: 99%

Advanced Techniques in Clinical Practice: Use of Lab-on-a-Chip Electrophoresis and Other Methods in Protein Profiling

Trenčevska¹,

Aleksovski²,

Стојаноски³

2009

Journal of Medical Biochemistry

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IntroductionEveryday laboratory analyses are done in order to contribute to precise medical diagnosis in patients with different diseases. The techniques developed include methods which are easy to handle, provide results with satisfying selectivity and sensitivity, and are fast enough to give results in a short time. Kratak sadr`aj: U klini~koj praksi, proteini se analiziraju kao parametri va`ni za odre|ivanje i le~enje razli~itih bolesti. Postoje dva opsega analiza: analize kompletnog proteinskog profila ili odre|ivanje izolovanog proteina. U ovom radu, pored kori{}enja uobi~ajenih metoda, pre svega elektroforeze, u analizu proteina uklju~ene su nove tehnike. Laboratorija na ~ipu je elektroforetska tehnika koja, opti mizovana, omogu}ava analize ukupnog proteinskog profila. Kada se normalni uzorci uporede sa uzorcima dobijenim od pacijenata s razli~itim neurolo{kim oboljenjima, postaju uo~lji vi karakteristi~ni obrasci. Tako|e, mogu}e je uspostaviti korelaciju i pore|enje izme|u nedavno razvijenih metoda »mikro~ip« elektroforeze i rezultata dobijenih kori{}enjem uobi~ajenih tehnika. Kada je neophodna ana liza specifi~nog proteina, masena spektrometrija je dala najbolje rezultate, {to se ti~e selektivnosti kao i specifi~nosti analiza. Postoji uverenje da je cistatin C potencijalni biomarker za neurolo{ka oboljenja, stoga je razvijen metod masene spektrometrije kako bi se po stigle kvalitativne i kvantitativne analize biolo{kih te~nosti. Pomo}u metode imunoeseja masenom spektrometrijom (MSIA), cistatin C je lako izolovan i analiziran a kompletna analiza trajala je nekoliko minuta. Maseni spektri koji su nastali kao rezultat otkrili su razli~ite nivoe izoformi cistatina C u serumu i uzorcima cerebrospinalne te~nosti. Summary: Proteins in clinical practice are analyzed as important parameters in the determination and treatment of different diseases. The scopes of the analyses are mainly concentrated in two levels -analyses of the complete protein profile, or determination of an isolated protein. In this work, despite of the use of conventional methods, mainly electrophoresis, new techniques have been implemented in protein analyses. Lab-on-a-chip is an electrophoretic techni que that, when optimized, provides analyses of the total protein profile. When normal samples are compared to samples obtained from patients with different neurological diseases, characteristic patterns can be noted. Also, correlation and comparison can be made between the newly developed microchip electrophoresis method and the results obtained using the conventional techniques. When an analysis of a specific protein is necessary, mass spectrometry has proven to give best results, in both the se lectivity and specificity of analyses. It is believed that cysta tin C is a potential biomarker in neurological diseases; the refore, the mass spectrometry method has been developed in order to obtain qualitative and quantitative analyses of biological fluids. Using the developed method of mass spectro metry immunoassay (MSIA), cystatin C was easily is...

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Targeted Mass Spectrometric Immunoassay for Detection of Cystatin C Isoforms in Cerebrospinal Fluid

Cited by 7 publications

References 12 publications

Multiplex Immuno-MALDI-TOF MS for Targeted Quantification of Protein Biomarkers and Their Proteoforms Related to Inflammation and Renal Dysfunction

Multiplex Immuno-MALDI-TOF MS for Targeted Quantification of Protein Biomarkers and Their Proteoforms Related to Inflammation and Renal Dysfunction

Developing Novel Methods for Protein Analysis and Their Potential Implementation in Diagnosing Neurological Diseases

Advanced Techniques in Clinical Practice: Use of Lab-on-a-Chip Electrophoresis and Other Methods in Protein Profiling

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