title: Factors influencing the measurement of lysosomal enzymes activity in human cerebrospinal fluid.
abstract: Measurements of the activities of lysosomal enzymes in cerebrospinal fluid have recently been proposed as putative biomarkers for Parkinson's disease and other synucleinopathies. To define the operating procedures useful for ensuring the reliability of these measurements, we analyzed several pre-analytical factors that may influence the activity of β-glucocerebrosidase, α-mannosidase, β-mannosidase, β-galactosidase, α-fucosidase, β-hexosaminidase, cathepsin D and cathepsin E in cerebrospinal fluid. Lysosomal enzyme activities were measured by well-established fluorimetric assays in a consecutive series of patients (n = 28) with different neurological conditions, including Parkinson's disease. The precision, pre-storage and storage conditions, and freeze/thaw cycles were evaluated. All of the assays showed within- and between-run variabilities below 10%. At -20°C, only cathepsin D was stable up to 40 weeks. At -80°C, the cathepsin D, cathepsin E, and β-mannosidase activities did not change significantly up to 40 weeks, while β-glucocerebrosidase activity was stable up to 32 weeks. The β-galactosidase and α-fucosidase activities significantly increased (+54.9±38.08% after 4 weeks and +88.94±36.19% after 16 weeks, respectively). Up to four freeze/thaw cycles did not significantly affect the activities of cathepsins D and E. The β-glucocerebrosidase activity showed a slight decrease (-14.6%) after two freeze/thaw cycles. The measurement of lysosomal enzyme activities in cerebrospinal fluid is reliable and reproducible if pre-analytical factors are accurately taken into consideration. Therefore, the analytical recommendations that ensue from this study may contribute to the establishment of actual values for the activities of cerebrospinal fluid lysosomal enzymes as putative biomarkers for Parkinson's disease and other neurodegenerative disorders.
authors.affiliation: Department of Pharmaceutical Sciences, University of Perugia, Perugia, Italy.
authors.forename: Emanuele
authors.initials: E
authors.lastname: Persichetti
authors.affiliation: Department of Medicine, Section of Neurology, University of Perugia, Perugia, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: Chiasserini
authors.affiliation: Department of Medicine, Section of Neurology, University of Perugia, Perugia, Italy.
authors.forename: Lucilla
authors.initials: L
authors.lastname: Parnetti
authors.affiliation: Regional Health Authority of Umbria, Epidemiology Department, Perugia, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Eusebi
authors.affiliation: Department of Pharmaceutical Sciences, University of Perugia, Perugia, Italy.
authors.forename: Silvia
authors.initials: S
authors.lastname: Paciotti
authors.affiliation: Department of Medicine, Section of Neurology, University of Perugia, Perugia, Italy.
authors.forename: Claudia
authors.initials: C
authors.lastname: De Carlo
authors.affiliation: Department of Pharmaceutical Sciences, University of Perugia, Perugia, Italy.
authors.forename: Michela
authors.initials: M
authors.lastname: Codini
authors.affiliation: Department of Medicine, Section of Neurology, University of Perugia, Perugia, Italy.
authors.forename: Nicola
authors.initials: N
authors.lastname: Tambasco
authors.affiliation: Department of Medicine, Section of Neurology, University of Perugia, Perugia, Italy.
authors.forename: Aroldo
authors.initials: A
authors.lastname: Rossi
authors.forename: Omar M A
authors.initials: OM
authors.lastname: El-Agnaf
authors.affiliation: Department of Biochemistry, Faculty of Medicine and Health Sciences, United Arab Emirates University, Al Ain, United Arab Emirates; Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia.
authors.forename: Omar M
authors.initials: OM
authors.lastname: El Agnaf
authors.affiliation: Department of Medicine, Section of Neurology, University of Perugia, Perugia, Italy; Fondazione S. Lucia, IRCCS, Rome, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Calabresi
authors.affiliation: Department of Pharmaceutical Sciences, University of Perugia, Perugia, Italy.
authors.forename: Tommaso
authors.initials: T
authors.lastname: Beccari
mesh_terms: D000368:Aged; D015415:Biomarkers / Q000134:cerebrospinal fluid; D002402:Cathepsin D / Q000134:cerebrospinal fluid; D020059:Cathepsin E / Q000134:cerebrospinal fluid; D005260:Female; D005962:Glucosylceramidase / Q000134:cerebrospinal fluid; D005966:Glucuronidase / Q000134:cerebrospinal fluid; D006801:Humans; D006867:Hydrolases / Q000134:cerebrospinal fluid*; D008247:Lysosomes / Q000201:enzymology*; D008297:Male; D008361:Mannosidases / Q000134:cerebrospinal fluid; D008875:Middle Aged; D010300:Parkinson Disease / Q000175:diagnosis / Q000201:enzymology*; D015203:Reproducibility of Results; D005644:alpha-L-Fucosidase / Q000134:cerebrospinal fluid; D043323:alpha-Mannosidase / Q000134:cerebrospinal fluid; D001616:beta-Galactosidase / Q000134:cerebrospinal fluid; D001619:beta-N-Acetylhexosaminidases / Q000134:cerebrospinal fluid
chemical_list: D015415:Biomarkers; D006867:Hydrolases; D008361:Mannosidases; D001616:beta-Galactosidase; D043323:alpha-Mannosidase; D005966:Glucuronidase; D005962:Glucosylceramidase; D005644:alpha-L-Fucosidase; D001619:beta-N-Acetylhexosaminidases; D020059:Cathepsin E; D002402:Cathepsin D