For histological analysis, osteochondral grafts were fixed in 4% buffered
formaldehyde solution (VWR, Radnor, PA, USA) for up to 1 week and decalcified
under constant agitation using Osteosoft solution (Merck, Burlington, MA, USA).
After decalcification (duration of 4-6 weeks), the osteochondral grafts were
embedded in Tissue-Tek OCT (optimal cutting temperature, VWR, Radnor, PA, USA)
and stored at −80°C. Sectioning was done using the CryoStar NX70 Cryostat
(Thermo Fischer Scientific, Waltham, MA, USA), with −25°C for the knife
temperature and −20°C for the chamber temperature. Six-micrometer sections were
obtained and processed for safranin O/light green staining. Images were taken
with a Leica DM-1000 microscope and processed using the Leica Manager software
(Leica, Wetzlar, Germany). To quantify changes within the cartilage sections
stained with safranin O/light green, a modified Mankin scoring system was used
(Table 3
The assessment was done by 5 independent observers with a maximum score
of 15.
formaldehyde solution (VWR, Radnor, PA, USA) for up to 1 week and decalcified
under constant agitation using Osteosoft solution (Merck, Burlington, MA, USA).
After decalcification (duration of 4-6 weeks), the osteochondral grafts were
embedded in Tissue-Tek OCT (optimal cutting temperature, VWR, Radnor, PA, USA)
and stored at −80°C. Sectioning was done using the CryoStar NX70 Cryostat
(Thermo Fischer Scientific, Waltham, MA, USA), with −25°C for the knife
temperature and −20°C for the chamber temperature. Six-micrometer sections were
obtained and processed for safranin O/light green staining. Images were taken
with a Leica DM-1000 microscope and processed using the Leica Manager software
(Leica, Wetzlar, Germany). To quantify changes within the cartilage sections
stained with safranin O/light green, a modified Mankin scoring system was used
(
The assessment was done by 5 independent observers with a maximum score
of 15.
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