Multivette 600

All animal studies were performed in accordance with protocols approved by the Institutional Animal Care and Use Committee at Roswell Park Cancer Institute. Male mice at 7-8 weeks of age were used, including wild type (WT) mice (C57BL/6), and mice deficient in FXII (C57BL/6-FXII^−/−) or FVII (C57BL/6-FVII^tTA/tTA). WT C57BL/6 mice were purchased from Taconic. C57BL/6-FXII^−/− mice and C57BL/6-FVII^tTA/tTA mice were bred in our own facility and genotyped as previously described [30 (link), 31 (link)]. The breeders were kindly provided by Dr. Francis J. Castellino at University of Notre Dame. All treatments were given by i.p. as follows: A single dose of vehicle, hPEPD, mPEPD, mSRC, hAβ1-42 or CCl₄; EP once daily for 5 days, followed at 1 h after the last dose of EP with a single dose of vehicle, hPEPD or hAβ1-42. CCl₄ was dissolved in corn oil, whereas all other substances were dissolved in PBS. The vehicle or the test substance was given to mice in 0.1 ml volume per 20 g body weight. Blood was collected from the mice at specific times by cardiac puncture at the time of sacrifice by carbon dioxide, and heart, liver and kidney were also collected from some of the mice. Blood was collected into K3 EDTA-containing tubes (Multivette 600 from Sarstedt), unless specified otherwise. All blood samples were promptly centrifuged to obtain plasma samples.

Immediately after blood sample collection into EDTA tubes (Multivette^® 600, Sarstedt Ag & Co.), 3 µL of whole blood was used to prepare a routine blood smear on single frosted glass slides. Three blood smears were prepared from each animal at each bleed. After air drying by waving slides in the air for few seconds, smears were fixed immediately in 100% methanol for 5 min. Smears were then air dried, before staining with Wright-Giemsa stain in a Siemens Hematek^® 1000 autostainer (Siemens Corp., Tarrytown, NY, USA). All blood smears were examined using a Nikon Eclipse Ci microscope at 10× or 20× objectives for manual cell counting and 20× to 40× objectives for differential counts.