Precise quantitation of 136 urinary proteins by LC/MRM-MS using stable isotope labeled peptides as internal standards for biomarker discovery and/or verification studies

Percy, Andrew J.; Yang, Juncong; Hardie, Darryl B.; Chambers, Andrew G.; Tamura-Wells, Jessica; Borchers, Christoph H.

doi:10.1016/j.ymeth.2015.04.001

Cited by 50 publications

(40 citation statements)

References 65 publications

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“…This panel of target proteins was selected from our in-house library of available SIS peptides that have been developed to target many potential biomarkers of clinically relevant conditions (i.e. lung cancer and cardiovascular disease) in various biofluids including cerebrospinal fluid, (33) plasma, (34,35) and urine (36). From this list, 68 proteins have been reported as FDA-cleared or -approved protein analytes in serum or plasma (see supplemental Table S2) (37).…”

Section: Methodsmentioning

confidence: 99%

Multiple Reaction Monitoring Enables Precise Quantification of 97 Proteins in Dried Blood Spots

Chambers

Percy

Yang

et al. 2015

Molecular & Cellular Proteomics

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The dried blood spot (DBS) methodology provides a minimally invasive approach to sample collection and enables room-temperature storage for most analytes. DBS samples have successfully been analyzed by liquid chromatography multiple reaction monitoring mass spectrometry (LC/MRM-MS) to quantify a large range of small molecule biomarkers and drugs; however, this strategy has only recently been explored for MS-based proteomics applications. Here we report the development of a highly multiplexed MRM assay to quantify endogenous proteins in human DBS samples. This assay uses matching stable isotope-labeled standard peptides for precise, relative quantification, and standard curves to characterize the analytical performance. A total of 169 peptides, corresponding to 97 proteins, were quantified in the final assay with an average linear dynamic range of 207-fold and an average R 2 value of 0.987. The total range of this assay spanned almost 5 orders of magnitude from serum albumin (P02768) at 18.0 mg/ml down to cholinesterase (P06276) at 190 ng/ml. The average intra-assay and interassay precision for 6 biological samples ranged from 6.1-7.5% CV and 9.5-11.0% CV, respectively. The majority of peptide targets were stable after 154 days at storage temperatures from ؊20°C to 37°C. Furthermore, protein concentration ratios between matching DBS and whole blood samples were largely constant (<20% CV) across six biological samples. This assay represents the highest multiplexing yet achieved for targeted protein quantification in DBS samples and is suitable for biomedical research applications. Molecular & Cellular Proteomics 14: 10.1074/mcp.O115.049957, 3094-3104, 2015.The dried blood spot (DBS) 1 methodology provides several advantages over traditional plasma or serum samples throughout the entire pre-analytical workflow including sample collection, transportation, and storage (1, 2) These blood samples are typically generated using a small sterile lancet to prick the skin and then spotting a drop onto a collection card. Therefore, DBS sampling is less invasive than venipuncture and does not require a trained phlebotomist. This sampling approach also does not require time-sensitive centrifugation, which is crucial for plasma and serum samples to prevent degradation. Many analytes have been determined to be stable in the DBS format at room temperature, eliminating the cost associated with cold-chain logistics for sample transportation and storage. These considerations are also important for sample collection in remote locations that may be without reliable access to a centrifuge and/or a freezer designated for biohazardous materials. Quantitative bioanalytical methods using the DBS methodology have been developed for genomic, metabolomic, and proteomic applications including newborn screening (3, 4), therapeutic drug monitoring (5, 6), toxicology and drugs of abuse (7,8), viral disease management (9, 10), and many others (2, 11).Targeted MS, in particular selected/multiple reaction monitoring (SRM/MRM) using internal standards...

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Section: Methodsmentioning

confidence: 99%

Multiple Reaction Monitoring Enables Precise Quantification of 97 Proteins in Dried Blood Spots

Chambers

Percy

Yang

et al. 2015

Molecular & Cellular Proteomics

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“…Biomarkers in the urine samples of infants are widely used for predicting such pathologies as obstructive nephropathy [11,12], necrotizing enterocolitis [13], and for monitoring maturation and development of the urinary system [7]. The majority of throughput urine proteome studies are focused on the adult population (control and disease) [14][15][16][17]. One of the first deep urine proteome studies initiated by the M. Mann's group resulted in the creation of the Max-Planck Unified (MAPU) proteome database which contains 1543 urinary proteins of adults [18].…”

Section: Introductionmentioning

confidence: 99%

“…Additional sample fractionation allowed Marimuthu and colleagues to add 671 proteins to this catalog [15]. Different strategies are applied for shotgun urinary proteomics, including semi-quantitative label-free HPLC-MS/MS, 2D-PAGE-MS/MS or CE-MS/MS [8][9][10][11][12][13]18], quantitative SRM/MRM HPLC-MS/MS [15][16][17].…”

Section: Introductionmentioning

confidence: 99%

Investigation of urine proteome of preterm newborns with respiratory pathologies

Стародубцева

Kononikhin

Бугрова

et al. 2016

Journal of Proteomics

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“…The protein HPR is more commonly found in the urine of patients with prostate cancer, though it has been found in one healthy individual in the study [29]. In another study, conducted by Percy et al described that 136 proteins, of greater than five order of magnitude, found in urine can be applied to prostate cancer samples, a possible means of diagnosis [30]. Bernardo et al further used the proteomic studies to gain insight on the pathogenesis of bladder cancer due to Schistosoma haematobium, which can also be used for early detection of urogenital schistosomiasis-induced bladder cancer [31].…”

Section: Proteomics In Urologymentioning

confidence: 84%

Future of Urological Diagnosis Using Proteomics: A Review of Literature

Dagur¹,

Jason²,

hi³

et al. 2016

Transl Biomed

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Proteomics is a large-scale study of proteins. Several technologies are used to isolate and identify proteins of various spectra, which is useful in differentiating complexity of proteins. Critical role of proteomics can be applied to the field of urology, as a means of noninvasive biomarkers to diagnose patients with benign and malignant nephro-urological diseases. The accuracy of proteomic analysis findings in predicting specific clinical pathologies continues to draw controversy, thus further studies are necessary to confirm the diagnosis. Future of diagnosis in urology is expected to be done by proteomic urinalysis.

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Precise quantitation of 136 urinary proteins by LC/MRM-MS using stable isotope labeled peptides as internal standards for biomarker discovery and/or verification studies

Cited by 50 publications

References 65 publications

Multiple Reaction Monitoring Enables Precise Quantification of 97 Proteins in Dried Blood Spots

Multiple Reaction Monitoring Enables Precise Quantification of 97 Proteins in Dried Blood Spots

Investigation of urine proteome of preterm newborns with respiratory pathologies

Future of Urological Diagnosis Using Proteomics: A Review of Literature

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