Peptide Charge Derivatization as a Tool for Early Detection of Preeclampsia by Mass Spectrometry—A Comparison with the ELISA Test

Abstract

Early detection of any preeclampsia biomarkers may lower the risk of mortality, both for a mother and a child. Our study focuses on techniques for preeclampsia biomarker identification by comparing the results of a method using liquid chromatography mass spectrometry in multiple reaction monitoring mode (LC-MS/MS) with those by the enzyme-linked immunosorbent assay (ELISA) test, as well as by comparing the obtained results with clinical data. In the proposed LC-MS/MS method a tryptic digest peptide charge derivatization strategy was used as a tool for sensitive detection of podocin, i.e., a previously discovered preeclampsia biomarker present in urine samples from pregnant women. Urine samples from pregnant women with diagnosed preeclampsia were collected at different stages of pregnancy and from healthy subjects, and then were analyzed by ELISA test and the proposed method with LC-MS/MS. Charge derivatization of the ε amino group of C-terminal lysine residues in tryptic digests by 2,4,6-triphenylpyrylium salt was performed to increase the ionization efficiency in the LC-MS/MS mode. Podocin was identified at the early stage of pregnancy, while its detection using an ELISA test was not possible. The protocol for urine sample preparation was optimized. Our results show that the proposed method by LC-MS/MS in combination with peptide charge derivatization, provides an ultrasensitive tool for diagnosis of preeclampsia, and provides earlier detection than a clinical diagnosis or ELISA test. The proposed solution may revolutionize medical diagnostics.

Qualitative and Quantitative Mass Spectrometry in Salivary Metabolomics and Proteomics

“…The last part of the reagent is the reactive group responsible for the selective reaction of the reagent with selected functional groups in the peptide [ 106 ]. The peptide derivatisation strategy proposed by us was applied in the proteomics investigation of the canine [ 108 ], equine [ 109 ], and human [ 110 , 111 ] urine samples, making their reliable and ultrasensitive analysis by mass spectrometry possible.…”

Catch, Modify and Analyze: Methods of Chemoselective Modification of Cysteine-Containing Peptides

“…We also proposed a technique to increase sensitivity and detectability of tryptic peptides—a charge derivatization with a quaternary ammonium tag in a the form of the 2,4,6-triphenypyrylium salt [ 44 , 48 , 57 ]. Furthermore, we compared and presented the advantage of the LC-MS method over the ELISA test in clinical diagnoses requiring identification of podocin [ 58 ].…”

“…However, all designed tags can be used for MS-based analyzes of hydrophobic peptides and low-abundance biomarkers. In the case of biomarker quantitation by MS, cysteine-containing peptides can be important candidates because of the stable ion generation by the derivatization of designed mass tags [ 58 , 62 ].…”

“…Furthermore, the proposed method can be extended to include reactivity toward other functional groups. ICAT reagents with different specificities could also make cysteine-free proteins susceptible to analysis by the ICAT method [ 58 ].…”

A Novel Bisquaternary Ammonium Compound as an Anion Sensor—ESI-MS and Fluorescence Study

“…This fragment can serve as a reporter ion in Multiple Reaction Monitoring analysis [ 18 ]. The proposed solution was successfully applied by us in the development of a method of human podocin tryptic peptide identification in urine sediment hydrolysates using mass spectrometry and chemical modification in the form of charge derivatization [ 19 ]. We also investigated the possibility of noncovalent dimer formation of a quaternary ammonium denatonium cation in the gas phase in ESI-MS analysis as a consequence of amide hydrogen deprotonation [ 20 ].…”