Absolute ProGRP quantification in a clinical relevant concentration range using LC–MS/MS and a comprehensive internal standard☆

Winther, Bjørn; Moi, Petter; Nordlund, Marianne S.; Lunder, Niclas; Paus, Elisabeth; Reubsaet, Léon

doi:10.1016/j.jchromb.2008.12.023

Cited by 21 publications

(13 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Optimal SRM conditions had been determined previously [12]. Repeatability, accuracy, linearity and LOD, LOQ were determined for the ProGRP signature peptide NLLGLIEAK in human serum after immuno-capture, in-well digestion and SPE.…”

Section: Evaluation Of the Methods 341 Methods Performancementioning

confidence: 99%

“…The analytical performance of the immuno-capture method was compared with the performance of the previously described LC-MS method utilizing protein precipitation and online sample cleanup (PPT-RAM method [12] Described by Nordlund et al [5], and regarded as reference method as it is used on routine basis for analyzing patient samples. Comparison with established immunofluorometric assay.…”

Section: Methods Comparisonmentioning

confidence: 99%

“…Concentrations down to 1.5 ng/mL (l200 fmol/mL) ProGRP in serum could be detected using acetonitrile (MeCN) induced protein precipitation and online sample cleanup prior to chromatography. Although clinically relevant, this LOD is still above the reference value [11,12]. Attempts to improve the LOD by using a triple quadropole mass spectrometer instead of a single quadropole mass spectrometer only contributed to a less complex chromatogram but not to a lower LOD.…”

Section: Introductionmentioning

confidence: 95%

“…This was discussed in ref. [12] and explained by possible ion-suppression, which might be circumvented by better sample preparation.…”

Section: Introductionmentioning

confidence: 99%

“…The low reference value of ProGRP has proven to be a challenge. Even optimized conventional sample preparation techniques, such as protein precipitation combined with online restricted access media (PPT-RAM), have fallen short with respect to sample cleanup and sensitivity [12]. The convenience of available antibodies for ProGRP [5] made it feasible to investigate immuno-capture as a sample cleanup step prior to tryptic digestion and LC-MS. A comparison of immuno-capture sample preparation and the PPT-RAM technique, applied to ProGRP, is performed with focus on sample cleanup, and effect on the LOQ.…”

Section: Introductionmentioning

confidence: 99%

See 4 more Smart Citations

Immuno‐capture as ultimate sample cleanup in LC‐MS/MS determination of the early stage biomarker ProGRP

Winther

Nordlund

Paus

et al. 2009

J of Separation Science

Self Cite

View full text Add to dashboard Cite

This paper presents a selective and efficient sample preparation procedure for NLLGLIEAK, signature peptide for the small cell lung cancer (SCLC) biomarker ProGRP, in human serum. The procedure is based on immuno-capture of ProGRP in 96-wells microtiter plates coated with the mAb E146. After immuno-capture and thorough rinse, trypsin was added for in-well digestion. Subsequently the signature peptide was enriched by SPE and determined by LC-MS/MS. Various steps in the procedure were optimized to achieve a low LOD such as dilution of sample, tryptic digestion, and SPE cleanup and peptide enrichment conditions. A single quadropole MS was used during optimization of the method. A triple quadropole MS was used in the method evaluation in order to improve sensitivity. The evaluation showed good repeatability (RSD, 11.9-17.5%), accuracy (3.0-6.6%), and linearity (r(2) = 0.995) in the tested range (0.5-50 ng/mL). LOD and LOQ were in the pg/mL area (0.20 and 0.33 ng/mL, respectively), enabling the determination of clinically relevant concentrations. The method was applied to two patient samples and showed good agreement with an established immunological reference method. The final method was compared to a previous published LC-MS method for the determination of ProGRP in serum based on protein precipitation and online sample cleanup. Both showed acceptable method performance, however, the immuno-capture LC-MS method was superior with respect to sensitivity. This illustrates the large potential of immuno-capture sample preparation prior to LC-MS in protein biomarker quantification.

show abstract

Section: Evaluation Of the Methods 341 Methods Performancementioning

confidence: 99%

Section: Methods Comparisonmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 95%

“…This was discussed in ref. [12] and explained by possible ion-suppression, which might be circumvented by better sample preparation.…”

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Immuno‐capture as ultimate sample cleanup in LC‐MS/MS determination of the early stage biomarker ProGRP

Winther

Nordlund

Paus

et al. 2009

J of Separation Science

Self Cite

View full text Add to dashboard Cite

show abstract

Current literature in mass spectrometry

2009

J. Mass Spectrom.

View full text Add to dashboard Cite

In order to keep subscribers up‐to‐date with the latest developments in their field, John Wiley & Sons are providing a current awareness service in each issue of the journal. The bibliography contains newly published material in the field of mass spectrometry. Each bibliography is divided into 11 sections: 1 Reviews; 2 Instrumental Techniques & Methods; 3 Gas Phase Ion Chemistry; 4 Biology/Biochemistry: Amino Acids, Peptides & Proteins; Carbohydrates; Lipids; Nucleic Acids; 5 Pharmacology/Toxicology; 6 Natural Products; 7 Analysis of Organic Compounds; 8 Analysis of Inorganics/Organometallics; 9 Surface Analysis; 10 Environmental Analysis; 11 Elemental Analysis. Within each section, articles are listed in alphabetical order with respect to author

show abstract

Clinical applications of MS‐based protein quantification

Sabbagh

Mindt

Neumaier

et al. 2016

Proteomics Clinical Apps

View full text Add to dashboard Cite

Mass spectrometry-based assays are increasingly important in clinical laboratory medicine and nowadays are already commonly used in several areas of routine diagnostics. These include therapeutic drug monitoring, toxicology, endocrinology, pediatrics, and microbiology. Accordingly, some of the most common analyses are therapeutic drug monitoring of immunosuppressants, vitamin D, steroids, newborn screening, and bacterial identification. However, MS-based quantification of peptides and proteins for routine diagnostic use is rather rare up to now despite excellent analytical specificity and good sensitivity. Here, we want to give an overview over current fit-for-purpose assays for MS-based protein quantification. Advantages as well as challenges of this approach will be discussed with focus on feasibility for routine diagnostic use.

show abstract

Absolute ProGRP quantification in a clinical relevant concentration range using LC–MS/MS and a comprehensive internal standard☆

Cited by 21 publications

References 18 publications

Immuno‐capture as ultimate sample cleanup in LC‐MS/MS determination of the early stage biomarker ProGRP

Immuno‐capture as ultimate sample cleanup in LC‐MS/MS determination of the early stage biomarker ProGRP

Current literature in mass spectrometry

Clinical applications of MS‐based protein quantification

Contact Info

Product

Resources

About