This protocol is adapted to determine total protein content using the Pierce BCA Protein Assay Kit from Thermo Scientific.

1. Thaw homogenized tissues
2. For 200 μL of coral larval tissues, add 10 μL of 1M NaOH (pH should be ~10) into each vial
   • The vial will be too small to fit a pH probe, therefore pipette a very small amount onto pH paper
3. Incubate each vial at 50°C for 4 hours shaking at 300rpm
4. Add 280 μL of 0.1M HCl to each vial to neutralize the sample
   • Add the HCl in small increments and test pH
   • pH should be at 7 - use more/less HCl depending on your sample
   • Ensure pH is at 7 before moving on to the next steps
5. Now use this prepared sample for the BCA protocol.

During the 4 hour incubation, follow the Pierce BCA Protein Assay Kit instructions to prepare the BSA standards and working reagents. Use the micro plate procedure if using the plate spectrophotometer.