picolorlock gold colorimetric assay reagents  (Thermo Fisher)

Journal: Molecular neurobiology

Article Title: Photobiomodulation for Global Cerebral Ischemia: Targeting Mitochondrial Dynamics and Functions

doi: 10.1007/s12035-018-1191-9

Figure Lengend Snippet: A–C, Western blotting analyses of Drp1 phosphorylation (Ser616, Ser637) and mitochondrial location of total Drp1, at ischemic reperfusion day 3 after GCI, using mitochondrial protein samples from CA1 subregion. D, Drp1 GTPase activity in hippocampal CA1 protein samples was quantified using a PiColorLock Gold kit. E, Total mitochondrial fragmentation in CA1 pyramidal neurons from sham, vehicle (Veh) and Mdivi-1 treated animals was examined 3 d after GCI. F, Typical staining of NeuN in vehicle (a) and Mdivi-1 (b) treated animals at day 14 after GCI, and the quantitative analyses of the numbers of surviving neurons in hippocampal CA1 layer (c). Dashed line represents percent value in sham control. Scale bar: 50 μm. Data are presented as means ± SE, n = 4-5 in A-E, n = 8-12 in F. *P < 0.05 versus sham control, #P < 0.05 versus GCI or GCI + sham PBM group. $P < 0.05 versus vehicle treated group.

Article Snippet: The beads were washed four times with GTPase wash buffer (50 mM Tris-HCl, pH 7.5, 1.0 M NaCl, 10 mM MgCl2, 4 mM DTT, 1 mM PMSF) and incubated with 0.5 mM GTP in reaction buffer (50 mM Tris-HCl, pH 7.5, 50 mM NaCl, 20 mM EDTA, and 5 mM MgCl2) at 30° C for 1 h. The released free phosphate was measured using a PiColorLock Gold Colorimetric Assay Kit (#303-0030, Novus Biologicals) according to the kit booklet protocol.

Techniques: Western Blot, Phospho-proteomics, Activity Assay, Staining, Control