After checking purity by HPLC analysis, metabolites were concentrated again by freeze-drying and re-suspended appropriately for structure determination.
Proteins in the active fraction were concentrated again via TCA precipitation (as before) and analysed by SDS page.
Pooled fractions were re-equilibrated in solution A without NP-40 (EconoPac 10 DG, BioRad) then concentrated in MSI Ultrafuge or Millipore (Bedford, MA) Ultrafree-4 centrifuge filters and mixed with a Macro-Prep High Q Support (BioRad) anion exchange resin in solution A. After centrifugation the preparations were concentrated again to less than 1 ml.
After removal of supernatant, the amoeba pellet was washed by addition of PBS, centrifuged again, and then placed in a flask with Bacto-casitone axenic medium and allowed to grow for another 7 10 days, after which the amoebas were concentrated again, washed, and placed into fresh axenic medium [ 22].
When the P2 fraction was further fractionated on a sucrose gradient, most of the acyltransferase activity was concentrated in the P3 fraction, again colocalizing with the ER marker protein calreticulin, but not with the Golgi marker TGN-46 (G4 and G3 fractions) (Figure 3C).
When n = 159, the zone of failure to reject the power law was concentrated around τ2 values of ∼2 5, again fairly insensitive to the values of τ3.
The "flowing hydrogel" could be formed again if the diluted solution was concentrated to above 0.5%.
The supernatant was concentrated using a rotary evaporator, dialyzed, and digested with Pronase again.
However, the surface area of the crater is a function of the area upon which the stresses are concentrated, which again depend on the morphology of the root surface as discussed earlier [21].
