Biomedical Engineering Reference
In-Depth Information
were streaked out on a Lysogeny broth medium (LB) agarose plate and incubated
over night at 37
◦
C. Single colonies were picked with a sterile needle and used to
inoculate a liquid LB culture that was incubated on a rotary shaker at 180 rpm at
37
◦
C. When the density of the culture reached an OD
600
of 0.1, 1ml of the bacterial
culture was centrifuged at maximum speed in an Eppendorf centrifuge for 5mins at
room temperature. The supernatant was discarded and the pellet was suspended in
1 ml of phosphate buffered saline (PBS), pH 7.0. For inactivation, the bacteria were
first heated to 75
◦
C for 15mins and then stored on ice.
2.3 Implant Preparation
For biocompatible and inflammatory implant preparation, respectively, plainmaterial
samples or samples coated with bacterial products were used. Porous implants were
used to increase the carrier capacity and the stability of the coatings to prolong the
release after implantation. Porous glass beads obtained from VitraPOR, Germany
(Size 4mm, Pore size-60
m) were used as biocompatible implants. Inflammatory
porous glass implants were prepared by soaking the beads for 2min in heat inactivated
Staphylococcus aureus
suspensions and left to dry under ambient conditions. Porous
titaniumdiscs of 7mmdiameter and 2mm thickness were prepared frommicro-beads
by an injection molding and sintering procedure. Magnesium discs with a diameter of
5mm and height of 2mmwere prepared by extrusion of a rod followed by cutting off
individual discs. Poly-L-lactic acid beads with a diameter of 5mm were purchased
from Good Fellow, England.
μ
2.4 Subcutaneous Implantations in Mice
Wild-type BALB/c mice were obtained from Harlan-Winkelmann laboratories,
Germany. Animals were housed under pathogen free conditions in a group of max-
imum five animals per cage. Mice were anesthetized by intraperitoneal injection of
ketamine (10mg/kg) and xylazine (4mg/kg). The back was shaved using an elec-
tronic razor (Aesculap, Germany). An incision of 1 cm was made in the dorsal skin
and a small pouch was made under the skin to insert the implant. The wound was
closed by interrupted suturing using polyglactin filaments (Ethicon, Germany). Mock
implantation was done by following the complete surgical procedure but without
inserting any implant. All animal experiments were done in accordance with the reg-
ulations and with the approval from the local authorities Lower Saxony State Office
for Consumer Protection and Food Safety (LAVES), permission number 33.42502/
07-10.5.
