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DECELLULARIZED AND FUNCTIONALIZED BOVINE GINGIVAL MATRIX,…

- - - - Change the solution every 24 hours.
        
        [After decellularization, dialyze the extracellular matrix (ECM) in the buffer solution for 24 hours.
        
        Change the solution every 6 hours to guarantee complete detergent removal.
        
        Place the fragments under agitation in deionized water for 72 hours changing the deionized water every 24 hours to eliminate any remaining residues.
- - - - For the reading, take a sample of the nanostructure from the original eppendorf and transfer it to a new eppendorf.
        
        Correctly identify the new eppendorfs and centrifuge them at 20°C and 5000 RPM for 10 minutes.
        
        After centrifugation, collect the supernatants and transfer them to a 96-well plate for reading.
        
        Perform the reading on the plate spectrophotometer at an absorbance of 204 nm.
        
        Repeat this procedure three times: after 30 minutes from the start of the process, after 6 hours, and after 24 hours.
- - - - After 24 hours, transfer the fragments to new wells.
        
        Prepare the Alamar Blue solution
        
        Add a sample of the Alamar Blue solution over the fragments and protect from light.
        
        Incubate in the incubator.
        
        Remove the supernatant and transfer to identified eppendorfs.
        
        Wash the fragments with the buffer solution and add a sample of medium. Incubate until the next day to repeat the Alamar Blue test.
        
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- - - - Histological inclusion
        
        Microtomy
        
        Histological staining
        
        Microscopy analysis and photo documentation