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However, future market needs will ultimately decide the final viability of mussel open ocean aquaculture.
With the standard PDT protocol, the curve progression is comparable but with a less intense effect and a final viability of 15.7%.
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As a result of this reduction limited the viability possible to achieve post-concentration, as the final viabilities were below the target by 15 20%.
Before the final cell viability assays, cells were cultured in doxorubicin-free medium for two weeks.
After the final wash, viability of cells was assessed using the 0.4% trypan blue dye exclusion method prior to and following transplantation.
Each sample was quantified in triplicate and averaged to obtain the final cell viability number (Supporting Information, Table S2).
However, the subsequent purification steps allowed refinement of the cell suspensions and the increase of the final cell viability.
Final product viability ranged between 96.03% and 99.97% (median 99.87%), exceeding the minimum threshold of 70% established in our previous experience [ 26] (see Table 1).
SCs were resuspended in 400 μl Annexin V binding buffer and incubated with 2 μl Annexin V-FITC at room temperature for 15 min, then 5 μg/ml (final concentration) viability dye propidium iodide was added.
According to the assay protocol, 25 µl of the MTT reagent (Sigma-Aldrich, MO, USA) was added to each well in order to assess cell viability (final concentration: 1 mg ml−1) at the end of the 24 h of incubation period.
To determine the optimal method for sputum homogenization different volumes of 0.1% dithiothreitrol (DTT) (Sigma, UK) were added to sputum samples, and the volume and rolling time determined by the sample's final macroscopic appearance, viability of the induced sputum mononuclear cells (trypan blue method), and the degree of background discoloration in the ELISPOT well.
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