Decrease in platelet, erythrocyte and lymphocyte acetylcholinesterase activities due to the presence of protease inhibitors in the storage buffer.
The effect of the presence of protease inhibitors in the storage buffer on acetylcholinesterase (AChE) activity was studied in crude membrane preparations from sheep platelets before and after solubilization of the membranes with Triton X-100. Although sensitive to the action of trypsin, the biological activity of AChE remained unchanged for as long as 6 days at 4 degrees C in a protease-inhibitor-free medium. At 10(-5) M final concentration PMSF reduced AChE activity to 50% after 24 hours of storage. This reduction was abolished in mixtures in which PMSF was present together with 3 mM EGTA, 5 mM EDTA and 0.1 mg/mL trypsin inhibitor. Nevertheless, under these conditions, storage periods longer than 24 hours still drastically reduced AChE activity. A mixture of EDTA, EGTA and trypsin inhibitor also produced a decrease in AChE activity after 24 hours. A more complex cocktail of inhibitors including several commonly used peptides decreased AChE activity only if PMSF or EDTA were present in the mixtures. Similar results were obtained with sheep erythrocytes or lymphocytes, and bovine erythrocytes.
Martín-Valmaseda
E M
EM
Department of Biochemistry and Molecular Biology, University of Salamanca, Spain.
Sánchez-Yagüe
J
J
Marcos
R
R
Llanillo
M
M
D000110:Acetylcholinesterase / Q000097:blood* / Q000187:drug effects*; D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology*; D001793:Blood Preservation; D002021:Buffers; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D004912:Erythrocytes / Q000187:drug effects / Q000201:enzymology*; D008214:Lymphocytes / Q000187:drug effects / Q000201:enzymology*; D011480:Protease Inhibitors / Q000494:pharmacology*; D012756:Sheep; D013696:Temperature; D013997:Time Factors
D002021:Buffers; D011480:Protease Inhibitors; D000110:Acetylcholinesterase