syntaxin-bp1 antibody (6d1)  (Bio-Techne corporation)

Journal: F1000Research

Article Title: A guide to selecting high-performing antibodies for STXBP1 (UniProt ID: P61764 ) for use in western blot, immunoprecipitation, and immunofluorescence

doi: 10.12688/f1000research.160174.1

Figure Lengend Snippet: Lysates of U-87 MG WT and STXBP1 KO were prepared, and 40 μg of protein were processed for western blot with the indicated STXBP1 antibodies. The Ponceau stained transfers of each blot are presented to show equal loading of WT and KO lysates and protein transfer efficiency from the acrylamide gels to the nitrocellulose membrane. Antibody dilutions were chosen according to the recommendations of the antibody supplier. Antibody dilution used: ab109023** at 1/1000, ab124920** at 1/10 000, ab315893** at 1/1000, A5420 at 1/1000, NBP3-15097** at 1/2000, AF5675 at 1/200, 13414** at 1/1000, GTX114808 at 1/500, GTX114809 at 1/500, 67137-1-Ig* at 1/5000, MA5-35796** at 1/2000, PA1-742 at 1/500 (2 μg/ml). Predicted band size: 67.5 kDa. **Recombinant antibody, *Monoclonal antibody.

Article Snippet: Bio-Techne (Novus Biologicals) , NBP3-15097 , 230461 , AB_3094817 , recombinant mono , S05-4B6 , rabbit , n/a , Wb, IF.

Techniques: Western Blot, Staining, Membrane, Recombinant

Journal: F1000Research

Article Title: A guide to selecting high-performing antibodies for STXBP1 (UniProt ID: P61764 ) for use in western blot, immunoprecipitation, and immunofluorescence

doi: 10.12688/f1000research.160174.1

Figure Lengend Snippet: U-87 MG lysates were prepared, and immunoprecipitation was performed using 0.6 mg of lysate and 2.0 μg of the indicated STXBP1 antibodies pre-coupled to Dynabeads protein A or protein G. Samples were washed and processed for western blot with the indicated STXBP1 antibody. For western blot, NBP3-15097** was used at 1/2000. The Ponceau stained transfers of each blot are shown. SM=4% starting material; UB=4% unbound fraction; IP=immunoprecipitate, HC= antibody heavy chain. **Recombinant antibody, *Monoclonal antibody.

Article Snippet: Bio-Techne (Novus Biologicals) , NBP3-15097 , 230461 , AB_3094817 , recombinant mono , S05-4B6 , rabbit , n/a , Wb, IF.

Techniques: Immunoprecipitation, Western Blot, Staining, Recombinant

Journal: F1000Research

Article Title: A guide to selecting high-performing antibodies for STXBP1 (UniProt ID: P61764 ) for use in western blot, immunoprecipitation, and immunofluorescence

doi: 10.12688/f1000research.160174.1

Figure Lengend Snippet: U-87 MG WT and STXBP1 KO cells were labelled with a green or a far-red fluorescent dye, respectively. WT and KO cells were mixed and plated to a 1:1 ratio on coverslips. Cells were stained with the indicatedSTXBP1 antibodies and with the corresponding Alexa-fluor 555 coupled secondary antibody including DAPI. Acquisition of the blue (nucleus-DAPI), green (WT), red (antibody staining) and far-red (KO) channels was performed. Representative images of the merged blue and red (grayscale) channels are shown. WT and KO cells are outlined with green and magenta dashed line, respectively. When an antibody was recommended for immunofluorescence by the supplier, we tested it at the recommended dilution. The rest of the antibodies were tested at 1 and 2 μg/ml and the final concentration was selected based on the detection range of the microscope used and a quantitative analysis not shown here. Antibody dilution used: ab109023** at 1/100, ab124920** at 1/100, ab315893** at 1/500, A5420 at 1/50, AF5675 at 1/200, NBP3-15097** at 1/500, 13414** at 1/100, GTX114808 at 1/800, GTX114809 at 1/1000, 67137-1-Ig* at 1/2000, MA5-35796** at 1/100, PA1-742 at 1/1000. Bars = 10 μm. **Recombinant antibody, *Monoclonal antibody.

Article Snippet: Bio-Techne (Novus Biologicals) , NBP3-15097 , 230461 , AB_3094817 , recombinant mono , S05-4B6 , rabbit , n/a , Wb, IF.

Techniques: Staining, Immunofluorescence, Concentration Assay, Microscopy, Recombinant

Journal: Acta Neuropathologica Communications

Article Title: MUNC18–1 gene abnormalities are involved in neurodevelopmental disorders through defective cortical architecture during brain development

doi: 10.1186/s40478-017-0498-5

Figure Lengend Snippet: Effects of Munc18–1 knockdown on subcellular distribution of N-Cadherin. a Localization of exogenous N-Cadherin in Munc18–1-deficient migrating neurons. E14.5 cerebral cortices were electroporated with pCAG-RFP plus pCAG-HA-N-Cadherin together with pSuper-H1.shLuc (i) or sh-Munc#1 (ii, iii) . Coronal sections were prepared at E18.0 and immunostained for HA-tag (i, ii) or HA-tag plus GM130 (iii) . (c) . Bars in ( i - ii ), 10 μm and ( iii ), 5 μm. b Quantification of N-Cadherin accumulation at Golgi. The ratio of RFP-positive cells with the accumulation was calculated for migrating neurons in the lower CP in ( a ). Error bars indicate SD; Control (n = 5), Munc18–1-knockdown (n = 5); ** p < 0.01 by Tukey-Kramer LSD. c Quantification of subcellular localization of N-Cadherin. The ratio of N-Cadherin in perinuclear to other cytoplasmic regions was analyzed. Error bars indicate SD of 5 brains containing more than 200 cells. ** p < 0.01 by Student’s t -test. d Localization of endogenous N-Cadherin in Munc18–1-deficient cortical neurons. pCAG-GFP was coelectroporated with pSuper-H1.shLuc (Control) or sh-Munc#1 into the E14.5 cerebral cortices. Neurons were isolated at E16.5, cultured for 48 h, fixed and stained with polyclonal anti-N-Cadherin without permeabilization. Bar, 10 μm. e Quantification of fluorescence intensity profiles of cell surface N-Cadherin across the cell bodies of control (blue) and Munc18–1-deficient neurons (red). Means +/− SEM (Control, n = 6; sh-Munc#1, n = 7). f Quantification of fluorescence intensity profiles of cell surface N-Cadherin in neurites. After the staining as in ( d ), neurons were permeabilized and double-stained with monoclonal anti-N-Cadherin and anti-GFP. Then, the ratio of the fluorescent intensity of surface N-Cadherin to total N-Cadherin was analyzed. Error bars indicate SD in each condition (n = 4). More than 300 neurites were analyzed in each experiment. ** p < 0.01 by Student’s t -test

Article Snippet: Mouse monoclonal anti-Munc18–1, anti-Syntaxin1A, anti-nestin and anti-N-Cadherin antibodies were purchased from Abnova Inc. (H00006812-M01; Taipei, Taiwan), Synaptic Systems (110,111; Gottingen, Germany), R&D Systems (MAB2736; Minneapolis, MN) and BD Biosciences (610,920; San Jose, CA), respectively.

Techniques: Knockdown, Control, Isolation, Cell Culture, Staining, Fluorescence