CM was then collected, concentrated using Agilent Concentrator (Agilent Technologies Inc). and filtered through 0.22 μM Millipore filter.
Extracellular media was collected, concentrated through 100,000 MWCO concentrators (Sartorius), loaded onto 10-60% Iodixanol gradients, and centrifuged at 100,000 g for 16 hours at 4°C.
Fractions containing absorbance peaks were collected, concentrated (∼5×) using Vivaspin 6 mL concentrators (Satorius, Goettingen, Germany) and run on SDS-PAGE and stained with Coomassie Brilliant Blue.
The conditioned medium was collected, concentrated with a Centricon-10 (UFC910024, Billerica, MA), and protein concentrations were determined with the BCA assay (#23250, Pierce, Rockford, IL).
Eighty (50 mL) fractions were collected, concentrated and grouped based on TLC profile.
The first peak was collected, concentrated to 5 mg/mL, and screened for crystallization.
Sixty (50 mL) fractions were collected, concentrated and grouped based on TLC profile.
The enzyme fractions were collected, concentrated, and exchanged with 10 mM phosphate buffer (pH 7.0).
The enzyme was collected, concentrated and partially purified from the culture supernatant.
The peaks in the elution profiles with purity above 95% were collected, concentrated by PEG20000 and stored at −80°C.
