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Absolute Quantification of Aβ1-42 in CSF Using a Mass Spectrometric Reference Measurement Procedure
Published on: March 21, 2017
Quantification of CSF cystatin C using liquid chromatography tandem mass spectrometry
Chikashi Matsuda1, Yuri Shiota1, Abdullah Md Sheikh2
1Clinical Laboratory Division, Shimane University Hospital, Japan.
Background:
Cystatin C (CST3), a ubiquitously expressed cysteine protease inhibitor, is implicated in several neurological diseases. Here, we have developed an accurate CST3 measurement system based on liquid chromatography tandem mass spectrometry (LC-MS/MS).
Methods:
LC-MS/MS based measurement for CSF CST3 was validated by determination of assay precision, accuracy and recovery. The values were compared with those measured by immunoassay. Glycosylation of CST3 in CSF was analyzed by Western blotting and lectin blotting.
Results:
Measuring standard CST3 by LC-MS/MS produced a linear standard curve that correlated with assigned values (r2=0.99). Both intra- and inter-assay variation was <10%. Although showed a correlation, the average CST3 concentration measured by LC-MS/MS was significantly higher than that of immunoassay. Western blotting showed the presence of a 25KDa species along with CST3 monomer (14KDa) in CSF. The volume of 25KDa species was decreased by deglycosylation. Lectin blotting revealed a 25KDa glycosylated protein in sialidase-treated CSF, which was decreased by deglycosylation. However, deglycosylation did not alter CST3 concentration measured by immunoassay.
Conclusions:
Our results suggest that LC-MS/MS-based CST3 measurement is a robust method with higher detection ability. Such method could be useful for the diagnosis and monitoring of neurological diseases.
