Journal: European Journal of Histochemistry : EJH

Article Title: Immunohistochemical Demonstration of Specific Antigens in the Human Brain Fixed in Zinc-ethanol-Formaldehyde

doi: 10.4081/ejh.2015.2530

Figure Lengend Snippet: Characteristics of the antibodies used and details of their application

Article Snippet: α-Tubulin , Mouse monoclonal (clone DM-1A); BioGenex, Fremont, CA, USA; MU121-UC; 1:100 , 60 min, 40°C , MACH2.

Techniques: Incubation

Journal: European Journal of Histochemistry : EJH

Article Title: Immunohistochemical Demonstration of Specific Antigens in the Human Brain Fixed in Zinc-ethanol-Formaldehyde

doi: 10.4081/ejh.2015.2530

Figure Lengend Snippet: Examples of immunohistochemical staining of the human brain fixed in the ZEF. A) Substantia nigra, pars reticulata : α-tubulin in the single neuron with extensively stained perikaryon and processes, many immunoreactive fibers in the surrounding neuropil. B) Nucleus ruber: extensive neuron-specific enolase (NSE) immunoreactivity in the nucleus, perinuclear cytoplasm, and processes of neurons; some bright NSE-immunoreactive fibers and medium-intensity staining of the neuropil are also seen. C) Caudate nucleus (head): choline acetyltransferase (ChAT) immunoreactivity in the multipolar neuron and its processes. D) Thoracic sympathetic ganglion: tyrosine hydroxylase (rabbit polyclonal) immunoreactivity in most of large and small neurons, mainly in the cytoplasm, as well as in bundles of nerve fibers going alongside these neurons. E) Substantia nigra, pars compacta : tyrosine hydroxylase (mouse monoclonal) antibody labels many (but not all) nigral neurons. F) Nucleus ruber: NeuN-labeled neurons with highly stained nucleus and the perinuclear cytoplasm. G) Parietal cortex, layer III: Iba-1 distinctly labels multiple thin processes and some perikarya of microgliocytes. H) Cerebellum: strong calbindin immunostaining in the cytoplasm and dendrites of the Purkinje cells. I) Cerebellum: calretinin immunohistochemistry visualizes the unipolar brush cell, a rare sort of the large cerebellar interneurons in the granular layer. J) S ubstantia nigra, pars reticulata : distinct synaptophysin immunoreactivity in the clusters of synapses alongside the neuronal and fiber plasmalemma. K) Cerebellar cortex: GAD65 immunostaining of the fine fiber nets in the granular layer, which localization coincides with the distribution of the glomeruli, the specific synaptic formations composed by the granular cell dendrites, mossy fibers, and axons of the Golgi cells. L) Nucleus ruber: GFAP distinctly labels the perinuclear cytoplasm and processes of fibrous astrocytes forming large nets in the neuropil. Preparations A-E), G-L) are counterstained with hematoxylin; F) with astra blue.

Article Snippet: α-Tubulin , Mouse monoclonal (clone DM-1A); BioGenex, Fremont, CA, USA; MU121-UC; 1:100 , 60 min, 40°C , MACH2.

Techniques: Immunohistochemical staining, Staining, Labeling, Immunostaining, Immunohistochemistry

Journal: The Journal of Biological Chemistry

Article Title: The PX-RICS-14-3-3?/? Complex Couples N-cadherin-?-Catenin with Dynein-Dynactin to Mediate Its Export from the Endoplasmic Reticulum *

doi: 10.1074/jbc.M109.081315

Figure Lengend Snippet: 14-3-3ζ/θ is involved in ER-to-Golgi transport of the N-cadherin-β-catenin complex. A, knockdown of 14-3-3ζ or -θ results in the disappearance of N-cadherin and β-catenin at the cell-cell boundaries and the ER accumulation of N-cadherin. HeLa cells transfected with the indicated siRNAs were subjected to immunofluorescent staining with anti-N-cadherin plus anti-calnexin or anti-β-catenin antibody. Arrows indicate N-cadherin or β-catenin enriched at the sites of cell-cell contact. Arrowheads indicate the absence of N-cadherin and β-catenin at the borders between two neighboring cells. Scale bars, 10 μm. B, upper panel, knockdown of 14-3-3ζ or -θ inhibits the surface expression of N-cadherin. The amounts of total and surface N-cadherin in siRNA-transfected HeLa cells were evaluated by surface biotinylation assays. Transferrin receptor (TfR) and α-tubulin were used for positive and negative controls, respectively. Lower panel, surface expression of N-cadherin was quantified by measuring the band intensity of the biotinylated fraction (SA pulldown in the upper panel) compared with that of total input. Representative results of the three independent experiments are shown. C, Ca2+-dependent cell adhesion is abrogated by knockdown of 14-3-3ζ or -θ. HeLa cells expressing the indicated siRNAs were dissociated by pipetting in the presence of Ca2+. Cell adhesion activity was quantified by counting the total number of cells (Nc) and the number of particles (cell clumps) (Np). Error bars represent the mean ± S.D. (n = 6).

Article Snippet: Mouse anti-14-3-3ϵ, mouse anti-β-catenin, and mouse anti-p150 Glued antibodies were purchased from BD Biosciences; mouse anti-GFP antibody (Living colors A.v. monoclonal antibody (JL-8)) was from Invitrogen; rabbit anti-Myc antibody was from MBL; mouse anti-α-tubulin antibody (DM1A) was from Merck; mouse anti-N-cadherin (13A9) and mouse anti-DYNC1I (74.1) antibodies were from Millipore; mouse anti-Myc (9E10), mouse anti-pan-14-3-3 (H-8), rabbit anti-14-3-3ζ (C-16), goat anti-14-3-3β (A-15), goat anti-14-3-3γ (A-12), goat anti-14-3-3η (E-12), and goat anti-Sec23 (E-19) antibodies were from Santa Cruz Biotechnology; mouse anti-FLAG (M2) and mouse anti-14-3-3θ (3B9) antibodies were from Sigma; rabbit anti-calnexin antibody was from Stressgen; and mouse anti-transferrin receptor antibody (H68.4) was from Zymed Laboratories Inc.. Rabbit and mouse IgGs were from Millipore.

Techniques: Transfection, Staining, Expressing, Activity Assay

Journal: bioRxiv

Article Title: LUZP1 regulates the constriction velocity of the contractile ring during cytokinesis

doi: 10.1101/2022.07.04.498656

Figure Lengend Snippet: (A) A plasmid encoding GFP-tagged full-length LUZP1 (GFP-LUZP1 FL) or GFP tag was transfected into HeLa cells. After 72 h, the cells were immunostained with anti-α- Tubulin and anti-GFP antibodies and Hoechst. The arrows indicate binucleated (multinucleated) cells (scale bar, 50 μm). (B) The ratio of multinucleated cells in (A) and (Supplemental Fig. S2A) was evaluated. Mean and SEM value of three independent experiments are shown; *, P < 0.05; n.s., not significant.

Article Snippet: Other antibodies and chemicals were obtained from the following manufacturers: anti- AIM-1 (AURKB) mouse monoclonal (6/AIM-1) antibodies (#611082) (BD Biosciences, Franklin Lakes, NJ, USA); anti-GFP polyclonal antibodies (#598) (Medical & Biological Laboratories, Tokyo, Japan); anti-α-Tubulin mouse monoclonal (DM1A) antibodies (#T6199) (Merck); anti-DAPK3 polyclonal antibodies (#PA5-90193) (Thermo Fisher Scientific, Waltham, MA, USA); anti-FLAG mouse monoclonal (M2) antibodies (#F3165) (Merck); Hoechst 33342 solution (Dojindo, Kumamoto, Japan); Nocodazole (Merck); Aurora kinase inhibitor VX-680 (also known as MK-0457, Tozasertib) (Merck); [γ- 32 P] ATP (PerkinElmer, Waltham, MA, USA); Trypsin Gold (Promega, Madison, WI, USA); Glutathione Sepharose 4B beads (GE Healthcare, Chicago, IL, USA); and reduced glutathione (Fujifilm).

Techniques: Plasmid Preparation, Transfection