After extensive washing with PBS buffer, PvNod41 that was bound to immobilized proteins on the matrix was recovered by boiling the sample with Laemmli buffer 2× [125 mM Tris-HCl pH 6.8, 4% SDS, 20% glycerol, 10% 2-mercaptoethanol, 0.02% (w/v) bromophenol blue] and analyzed by 12% SDS-PAGE and Coomassie Blue staining.
Capture antibody diluted in 50 mM carbonate buffer pH 9.6, was bound to polysorb 96-well EIA plates (Nunc, Denmark) overnight at 4°C.
The sample was then applied directly to a silica membrane-containing column and the RNA was bound and cleaned using buffers provided by the manufacturer to remove impurities.
As evidence of transplacental transfer of MSP142 occurring in the form of ICs, we noted that more MSP1 in fetal perfusate was detected following treatment with dissociation buffer, indicating that at least some MSP1 was bound to antibody (Figure 3).
In the NO-off form in vitro, either water or ammonia, depending on buffer type, is bound to the sixth site.
Beads were washed three times in RIPA buffer, and proteins that were bound to streptavidin beads were eluted in 1× Laemmli sample buffer containing 2-mercaptoethanol.
After incubation with gentle rocking overnight at 4°C, the beads were washed five times with lysis buffer, and proteins that were bound to the beads were analyzed by Western blot.
Aliquots (70 µl) of Dynabeads (Invitrogen, Carlsbad, CA), pre-coated with sheep anti-rabbit IgG, in buffered saline were bound with 6 µg affinity-purified rabbit antibodies to βI-spectrin or with purified non-immune rabbit IgG and washed.
In two of our DEAD-domain crystal structures, anions from the crystallization buffers are bound to motif Ib (a sulfate in DDX5, and a phosphate in DDX47) highlighting the ability of this motif to bind polyanions.
After ∼1 ms, the steady state, in which most of the stationary buffers were bound to Ca2+, was achieved.
