Development of an ultrasensitive enzyme immunoassay for the determination of matrix metalloproteinase-9 (MMP-9) levels in normal human cerebrospinal fluid.
Determination of matrix metalloproteinase-9 (MMP-9) in human cerebrospinal fluid (CSF) to study blood-brain barrier impairment and immune cell migration in inflammatory neurological diseases recently became a matter of major interest. Regularly, MMP-9 was determined qualitatively or semi-quantitatively by zymography (gelatin gel electrophoresis) or quantitatively by enzyme immunoassay (EIA). As yet, it was not possible by either method to detect MMP-9 in CSF of controls (patients without pathologically increased CSF parameters). We developed an ultrasensitive two-side enzyme-linked immunosorbent assay (ELISA) which allows for the first time to measure reliably MMP-9 concentrations in CSF of controls. This ELISA uses a monoclonal as capture and a polyclonal as detector antibody. The detection limit of the assay is below 10 pg/ml and the assay range is 15-2000 pg/ml. Intra-assay precision is 2.5% for low and 3.7% for high, inter-assay precision is 11% for low and 10.7% for high values, respectively. The determination of the MMP-9 concentration in 50 control CSF gave the following results: range, 22-146 pg/ml; median, 76 pg/ml. The measurement of native and recombinant MMP-9 was carried out with three commercially available ELISAs, most widely employed in MMP-9 research, and compared to the newly developed one. All ELISAs recognize recombinant MMP-9 by factors of 5-20 less sensitively than native MMP-9.
Maliszewska
M
M
Department of Neurology, Georg August University, Robert-Koch-Str. 40, D-37075, Göttingen, Germany.
Mäder
M
M
Schöll
U
U
Azeh
I
I
Hardeland
R
R
Felgenhauer
K
K
Beuche
W
W
Weber
F
F
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000592:standards; D005260:Female; D006801:Humans; D008297:Male; D020780:Matrix Metalloproteinase 9 / Q000032:analysis / Q000134:cerebrospinal fluid*; D008875:Middle Aged; D012016:Reference Values; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D020780:Matrix Metalloproteinase 9