Model 28000

Dressings and wound bed tissue were evaluated by scanning electron microscopy (SEM). A representative mouse from each of the three separate experiments in the placebo, 0.1% GaCi BID, and 0.3% GaCi OD treatment groups was sacrificed at 4 h postwounding and on day 7. The transparent dressings and a 4-mm tissue disc for each animal were fixed in 4% formaldehyde, 1% glutaraldehyde, and 0.1 M PBS. The samples were washed three times using 0.1 M PBS and then postfixed in 1% osmium tetroxide in 0.1 M PBS for 1 h. The samples were dehydrated in a graded series of ethanol solutions and then dried (critical point dryer, model 28000; Ladd Research Industries, Burlington, VT). The samples were mounted by double-sided carbon tape to specimen stubs and ion coated with gold/palladium (30:70) (Hummer X sputter coater; Anatech Ltd., Alexandria, VA). The samples were visualized using an Amray 3600 field emission (FE) scanning electron microscope (Bedford, MA) operated at a voltage of 3 kV. Samples were analyzed by scanning 10 or more fields at 1,000× magnification within the wounded tissue and on the portion of the dressing overlying the wounded area. Photomicrographs representative of the observed biofilm density were taken at 2,500× magnification.

A single wound was evaluated per pig on days 2, 4, 7, and 10 postinoculation. The transparent dressings and a 4 mm biopsy punch tissue sample for each animal were fixed in 4% formaldehyde, 1% glutaraldehyde, and 0.1 M PBS. The samples were washed three times using 0.1 M PBS and then postfixed in 1% osmium tetroxide in 0.1 M PBS for 1 h. The samples were dehydrated in a graded series of ethanol solutions and then dried (Critical point dryer, Model 28000; Ladd Research Industries, Burlington, VT). The samples were mounted using a double-sided carbon tape to specimen stubs. They were then ion coated with gold:palladium (30:70) (Hummer X Sputter Coater; Anatech Ltd., Alexandria, VA). The samples were then visualized using an Amray 3600 FE scanning electron microscope (Bedford, MA) operated at a voltage of 3 kV and analyzed by scanning 10 or more 1,000 × magnified fields within the wounded tissue and on the portion of the dressing overlying the wounded area. Photomicrographs representative of the observed bacterial density were taken at 2,500 × magnification, which appears to be a biofilm.