*4.3. Collagen and Protein Quantification*

Sircol Collagen Assay (Biocolor®, Carrick, UK) and BCA assay (MicroBCA protein detection kit, Pierce®, Thermo fisher Scientific, Waltam, MA, USA) were performed on the extracts from the choanosome according to the indications provided by the manufacturer, to quantify the amount of collagen and total proteins in the extracts, respectively. Readings were performed on a plate microreader (Synergy HT, Bio-Tek Instruments, Winooski, VT, USA), with calibration curve and concentrations of collagen/proteins being obtained by using the mean of at least three values for each standard/sample. Both Sircol and BCA assay were performed only on Ch hydrogels. The total amount of the extracted material was calculated by weighting standard amounts of freeze-dried material (mg/mL) and multiplying the values for the final volumes.
