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Image studio lite 3

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Image Studio Lite 3.1 is a software application designed for analyzing and quantifying images from various sources, including Western blots, electrophoresis gels, and other imaging techniques. The software provides basic image analysis tools and functionality.

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Lab products found in correlation

Bca protein assay, by Thermo Fisher Scientific (4 mentions) Ripa buffer, by Thermo Fisher Scientific (4 mentions) Irdye 800cw, by LI COR (3 mentions) Protease and phosphatase inhibitor cocktail, by Thermo Fisher Scientific (2 mentions) Mouse anti gapdh, by Thermo Fisher Scientific (2 mentions) Alexa fluor 680, by Thermo Fisher Scientific (2 mentions) Mouse anti defa5, by Santa Cruz Biotechnology (2 mentions) Goat anti actin, by Merck Group (2 mentions) Blocking buffer, by LI COR (2 mentions) Odyssey infrared imaging system, by LI COR (2 mentions) Laemmli sample buffer, by Bio-Rad (2 mentions) Prism 6, by GraphPad (1 mentions) Snap i d system, by Merck Group (1 mentions) Ab68718, by Abcam (1 mentions) Nb100 56875, by Novus Biologicals (1 mentions) Criterion tgx gel, by Bio-Rad (1 mentions) Bradford assay, by Bio-Rad (1 mentions) Rabbit anti gapdh, by Abcam (1 mentions) Irdye 680, by LI COR (1 mentions)

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Image Studio Lite version 3.1 Image Studio Lite Ver 3.1 Image Studio Lite Image Studio 3.1 software Image Studio

5 protocols using image studio lite 3

1

Crizotinib and PF-06463922 Inhibit ALK Signaling

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PC12 cells were grown and transfected with various ALK constructs as described (Chand et al., 2013 (link)). 48 h after transfection, cells were treated with different concentrations of crizotinib (from 0 to 5000 nM) or PF-06463922 (from 0 to 500 nM) for 2 h. Cells were then washed twice with PBS and harvested as described (Schonherr et al., 2012 (link), 2011 (link)). Samples were boiled in SDS sample buffer and analyzed by immunoblotting. Intensity of pALK (Y1604) and total ALK bands was quantified with Image Studio Lite 3.1 (LI-COR). Data were normalized to the 0 nM inhibitor samples. IC50 values were calculated by fitting data to a log (inhibitor) versus normalized response (variable slope) equation in GraphPad Prism 6.0.
Guan J., Tucker E.R., Wan H., Chand D., Danielson L.S., Ruuth K., El Wakil A., Witek B., Jamin Y., Umapathy G., Robinson S.P., Johnson T.W., Smeal T., Martinsson T., Chesler L., Palmer R.H, & Hallberg B. (2016). The ALK inhibitor PF-06463922 is effective as a single agent in neuroblastoma driven by expression of ALK and MYCN. Disease Models & Mechanisms, 9(9), 941-952.
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2

Western Blot Analysis of OX1R in Hippocampus

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Hippocampal tissue was micro-dissected and protein extraction was performed using a RIPA commercial protein extraction kit (Thermo Fisher Scientific, Rockford, IL) [33 (link)]. Protein quantification was determined using a Bradford assay (BioRad, Hercules, CA) and samples (10 μg per well) were separated on 10% Criterion TGX gel (BioRad) and then transferred to PVDF membrane, and blocked (Super Blocker; Thermo Fisher Scientific) using Snap ID system (Millipore, Billerica, MA). Antibodies: OX1R (ab68718, Abcam, Cambridge, MA) and GAPDH (NB100-56875; Novus) primary; HRP conjugated anti-rabbit IgG (NB710H, Novus, Littleton, CO) secondary. All primary antibodies were used at a 1:1000 dilution, while secondary antibodies were used at a final dilution of 1:60,000. Protein samples were visualized and band density was determined (LICOR Image Studio Lite 3.1, Lincoln, NE). Data were normalized to GAPDH [36 (link),37 (link)], and analyzed using unpaired t-test using GraphPad Prism.
Mavanji V., Butterick T.A., Duffy C.M., Nixon J.P., Billington C.J, & Kotz C.M. (2017). Orexin/hypocretin treatment restores hippocampal-dependent memory in orexin-deficient mice. Neurobiology of learning and memory, 146, 21-30.
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3

Western Blot Analysis of Cellular Proteins

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Cells were lysed in cold RIPA buffer (Thermo Fisher Scientific, Rockford, IL) supplemented with 1% protease and phosphatase inhibitor cocktail (Thermo Fisher Scientific). Protein concentrations were determined with the BCA protein assay (Thermo Fisher Scientific). Cell lysates were suspended in × Laemmli sample buffer (Bio-Rad, Hercules, CA) containing 5% 2-mercaptoethanol and boiled for 5 minutes. After heat denaturation, total protein lysates (30 ug/lane) were subjected to tricine-SDS-PAGE [25 ]. The proteins were then transferred electrophoretically to 0.2 um PVDF membranes. Membranes were blocked in blocking buffer (LI-COR Biosciences, Lincoln, NE) diluted 1:1 in × PBS for 1 hour at room temperature. The blots were incubated with mouse anti-DEFA5 (Santa Cruz Biotechnologies, Santa Cruz, CA; 1:200 dilution), goat anti-actin (Sigma; 1:1000 dilution), and mouse anti-GAPDH (Life Technologies; 1:1000 dilution) overnight at 4°C. After washing with PBST, blots were incubated with Alexa Fluor 680 (Life Technologies) and IRDye 800CW (LI-COR Biosciences) conjugated secondary antibodies (1:10,000 dilution) for 1 hour at room temperature. The band intensities were quantified using an Odyssey infrared imaging system (LI-COR Biosciences) and analyzed using Image Studio Lite 3.1 (LI-COR Biosciences).
Miles D.R., Shen J., Chuang A.Y., Dong F., Wu F, & Kwon J. (2016). Alpha-Defensin 5 Expression is Regulated by microRNAs in the Caco-2 Intestinal Epithelial Cell Line. Journal of inflammatory bowel diseases & disorders, 1(1), 105.
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4

Protein Isolation and Western Blot Analysis

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Protein isolation and western blot analysis were performed using standard protocols. We used the following primary antibodies: mouse anti-Shank2 (UC Davis/NIH NeuroMab Facility, Davis, CA, USA), rabbit anti-actin (Cytoskeleton, Denver, CO, USA) and rabbit anti-GAPDH (Abcam, Cambridge, UK). IRDye 800CW and IRDye 680 (LI-COR Biosciences, Lincoln, NE, USA) served as secondary antibodies. All western blot band quantification procedures were performed using Image Studio Lite 3.1 software (LI-COR Biosciences).
Peykov S., Berkel S., Schoen M., Weiss K., Degenhardt F., Strohmaier J., Weiss B., Proepper C., Schratt G., Nöthen M.M., Boeckers T.M., Rietschel M, & Rappold G.A. (2015). Identification and functional characterization of rare SHANK2 variants in schizophrenia. Molecular Psychiatry, 20(12), 1489-1498.
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5

Western Blot Analysis of Cellular Proteins

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Cells were lysed in cold RIPA buffer (Thermo Fisher Scientific, Rockford, IL) supplemented with 1% protease and phosphatase inhibitor cocktail (Thermo Fisher Scientific). Protein concentrations were determined with the BCA protein assay (Thermo Fisher Scientific). Cell lysates were suspended in × Laemmli sample buffer (Bio-Rad, Hercules, CA) containing 5% 2-mercaptoethanol and boiled for 5 minutes. After heat denaturation, total protein lysates (30 ug/lane) were subjected to tricine-SDS-PAGE [25 ]. The proteins were then transferred electrophoretically to 0.2 um PVDF membranes. Membranes were blocked in blocking buffer (LI-COR Biosciences, Lincoln, NE) diluted 1:1 in × PBS for 1 hour at room temperature. The blots were incubated with mouse anti-DEFA5 (Santa Cruz Biotechnologies, Santa Cruz, CA; 1:200 dilution), goat anti-actin (Sigma; 1:1000 dilution), and mouse anti-GAPDH (Life Technologies; 1:1000 dilution) overnight at 4°C. After washing with PBST, blots were incubated with Alexa Fluor 680 (Life Technologies) and IRDye 800CW (LI-COR Biosciences) conjugated secondary antibodies (1:10,000 dilution) for 1 hour at room temperature. The band intensities were quantified using an Odyssey infrared imaging system (LI-COR Biosciences) and analyzed using Image Studio Lite 3.1 (LI-COR Biosciences).
Miles D.R., Shen J., Chuang A.Y., Dong F., Wu F, & Kwon J. (2016). Alpha-Defensin 5 Expression is Regulated by microRNAs in the Caco-2 Intestinal Epithelial Cell Line. Journal of inflammatory bowel diseases & disorders, 1(1), 105.
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