Methodological issues affecting the determination of plasma matrix metalloproteinase (MMP)-2 and MMP-9 activities.
OBJECTIVE
To examine whether a linear relationship exists between the volume of plasma loaded in each gel lane and the gelatinolytic activities of plasma matrix metalloproteinase (pMMP)-2 and pMMP-9, and the influences of plasma storage temperature, fasting, and a variable number of repeated freeze-thaw cycles.


METHODS
Plasma from venous blood collected into EDTA tubes was stored for 1 month at -20 degrees C or at -70 degrees C until assayed for gelatin zymography of pMMP-2 and pMMP-9. The effects of food intake, repeated freeze-thaw cycles, and the gelatinolytic activity produced by serial dilutions of plasma were examined.


RESULTS
Storage temperature, fasting, and up to five repeated freeze-thaw cycles had no effect on the activities of pMMPs (all P < 0.05). A linear relationship was found with plasma volumes below 0.75 microL/lane for MMP-2, and from 0.75 microL/lane to 2 microL/lane for MMP-9.


CONCLUSIONS
Different volumes of plasma should be studied when pMMP-2 and pMMP-9 activities are studied. Storage temperature, repeated freeze-thaw cycles, and food intake do not affect the activities of pMMPs.
Souza-Tarla
Caroline D
CD
Department of Pharmacology, Faculty of Medicine of Ribeirao Preto, University of Sao Paulo, Av. Bandeirantes, 3900, 14049-900 Ribeirao Preto, SP, Brazil.
Uzuelli
Juliana A
JA
Machado
Alcyone A
AA
Gerlach
Raquel F
RF
Tanus-Santos
Jose E
JE
D001800:Blood Specimen Collection; D004435:Eating; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005615:Freezing; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000097:blood*; D020780:Matrix Metalloproteinase 9 / Q000097:blood*
D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9