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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Winther, Bjørn Nordlund, Marianne Paus, Elisabeth Reubsaet, Léon Halvorsen, Trine Grønhaug |
| Description | Country affiliation: Norway Author Affiliation: Winther B ( Department of Pharmaceutical Chemistry, School of Pharmacy, University of Oslo, Oslo, Norway.) |
| Abstract | 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. |
| File Format | HTM / HTML |
| ISSN | 16159306 |
| Issue Number | 17 |
| Volume Number | 32 |
| e-ISSN | 16159314 |
| Journal | Journal of Separation Science |
| Language | English |
| Publisher | Wiley-VCH |
| Publisher Date | 2009-09-01 |
| Publisher Place | Germany |
| Access Restriction | One Nation One Subscription (ONOS) |
| Subject Keyword | Discipline Analytical Discipline Chemistry Biological Markers Chromatography, Liquid Methods Immunoassay Peptide Fragments Tandem Mass Spectrometry Blood Chemistry Humans Instrumentation Genetics Recombinant Proteins Reproducibility Of Results Sensitivity And Specificity Small Cell Lung Carcinoma Metabolism Evaluation Studies Journal Article |
| Content Type | Text |
| Resource Type | Article |
| Subject | Analytical Chemistry Filtration and Separation |
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