Capillary electrophoresis with laser-induced fluorescence detection as a novel sensitive approach for the analysis of desmosines in real samples.
Among various biomarkers believed to behave as descriptors of the disease process in chronic obstructive pulmonary disease (COPD), urinary desmosines are commonly used for monitoring elastin degradation. Given the low concentrations of urinary desmosines, their quantitative determination in this biological matrix often requires preconcentration steps. To minimize both solute losses and effects of sample matrix, and to decrease data variability related to the above-mentioned manipulation processes, we have developed a capillary electrophoresis approach combined with laser-induced fluorescence (CE-LIF) detection system using urine samples not submitted to any pretreatment procedure other than filtering the sample. Urines were hydrolyzed, derivatized with fluorescein isothiocyanate (FITC) and endogenous desmosines were identified by addition of standard analytes and submitting to mass spectrometry (MS) analysis the material collected from micropreparative runs. The assay showed good linearity, reproducibility and precision, allowing to detect amounts of desmosines as low as 10(-8) M (equivalent to 0.1 fmol on column). We conclude that CE-LIF technique is a highly sensitive method for detecting urinary desmosines.
Dipartimento di Biochimica A Castellani, Università di Pavia, Italy.
Laura
L
Annovazzi
Simona
S
Viglio
Eleonora
E
Perani
Maurizio
M
Luisetti
James
J
Baraniuk
Begoña
B
Casado
Giuseppe
G
Cetta
Paolo
P
Iadarola
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D003895:Desmosine / Q000652:urine*; D019075:Electrophoresis, Capillary / Q000379:methods*; D016650:Fluorescein-5-isothiocyanate; D005453:Fluorescence; D006801:Humans; D007834:Lasers; D013058:Mass Spectrometry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet; D013995:Time
D015415:Biomarkers; D003895:Desmosine; D016650:Fluorescein-5-isothiocyanate