Image studio v4

Manufactured by LI COR

Sourced in United States

Image Studio v4.0 is a software application designed to analyze and quantify images generated by various imaging devices. It provides tools for image processing, data analysis, and visualization. The software is compatible with a range of imaging platforms and can be used to measure and interpret signals from various experimental techniques.

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Lab products found in correlation

Ripa buffer, by Thermo Fisher Scientific (4 mentions) Odyssey system, by LI COR (3 mentions) Ecl plus, by Thermo Fisher Scientific (3 mentions) Protease inhibitor, by Merck Group (2 mentions) Intercept tbs blocking buffer, by LI COR (2 mentions) Odyssey clx imaging system, by LI COR (2 mentions) Bca protein assay, by Thermo Fisher Scientific (2 mentions) Gradient sds page, by Bio-Rad (2 mentions) Ripa buffer, by Teknova (2 mentions) Odyssey fc imaging system, by LI COR (2 mentions) Dc protein assay, by Bio-Rad (2 mentions) Protease inhibitor cocktail, by Roche (2 mentions) Odyssey blocking buffer, by LI COR (2 mentions) Odyssey, by LI COR (2 mentions) Irdye 680rd, by LI COR (2 mentions) Nupage 4 12 bis tris gel, by Thermo Fisher Scientific (2 mentions) Irdye 800, by LI COR (2 mentions) Nitrocellulose membrane, by Bio-Rad (2 mentions) Odyssey clx system, by LI COR (1 mentions) Alexa fluor 680 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)

16 protocols using image studio v4

Western Blot Analysis of Viral Proteins

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Cells collected for western blot analysis were lysed in lysis buffer. Protein concentration was measured by Pierce BCA assay (ThermoFisher). Ten μg were reduced with 2-mercaptoethanol and subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane, and the membranes were blocked in blocking buffer (5% milk in TBS), primary antibody in blocking buffer with 0.2% Tween, and secondary antibody in blocking with 0.2% Tween and 0.01% SDS. Fluorescent signal from secondary antibodies was detected using the Odyssey CLX system (Li-cor). ImageStudio v4.0 (Li-cor) was used for band intensity quantification.
The following antibody dilutions were used for primary incubation: 1:200 for mouse anti-pp28 (clone 10B4 gift from Tom Shenk (Silva et al., 2003 (link))), 1:100 for mouse anti-IE1 (clone 1B12 gift from Tom Shenk (Zhu et al., 1995 (link))), 1:100 for mouse anti-pUL26 (clone 7H1-5 gift from Tom Shenk (Munger et al., 2006 (link))), 1:100 for mouse anti-pp65 (clone 8F5 (Nowak et al., 1984 (link))), 1:5000 for mouse anti-α-tubulin (ThermoFisher T6199), 1:2000 for rabbit anti-PEX14 (Abcam ab183885), 1:500 rabbit anti-GNPAT (Proteintech 14931-1-AP). The following antibodies at 1:10000 dilutions were used for secondary incubations: Alexa Fluor 680 goat anti-rabbit IgG (ThermoFisher A-21109) and IRDye 800CW goat anti-mouse (Li-cor 926-32210).

Jean Beltran P.M., Cook K.C., Hashimoto Y., Galitzine C., Murray L.A., Vitek O, & Cristea I.M. (2018). Infection-induced peroxisome biogenesis is a metabolic strategy for herpesvirus replication. Cell host & microbe, 24(4), 526-541.e7.

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Western Blot Analysis of Abcg5 and Abcg8

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Liver and intestines were lysed by using lysis buffer and underwent western blot analysis with the primary antibodies anti-Abcg5 (1:1000; sc-25796, Santa Cruz, CA, United States), anti-Abcg8 (1:1000; sc-30111, Santa Cruz, CA, United States), and β-tubulin (1:1000; 200608, Zen Bio Science, China). Bands were visualized by using the LI-COR (Lincoln, NE, United States) Odyssey System. Quantification of band intensity involved using Image Studio v4.0 (LI-COR).

Li H., Shen J., Wu T., Kuang J., Liu Q., Cheng S., Pu S., Chen L., Li R., Li Y., Zou M., Zhang Z., Jiang W., Qu A, & He J. (2019). Irisin Is Controlled by Farnesoid X Receptor and Regulates Cholesterol Homeostasis. Frontiers in Pharmacology, 10, 548.

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Protein Isolation and Immunoblot Analysis

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Total proteins were isolated using RIPA Buffer (Thermo Scientific) and freeze-thawing at –80°C in the presence of 1× Halt Protease Inhibitor (Thermo Scientific), 5 mM ethylenediaminetetraacetic acid (EDTA), and Phosphatase Inhibitor Cocktails 2 and 3 (Sigma-Aldrich). Protein concentration was determined using a BCA Protein Assay (Thermo Scientific), and 50 μg protein was separated via electrophoresis on a Bolt 4–12% Bis-Tris Plus Gel (Life Technologies) and transferred onto membranes using the Iblot2 (Life Technologies) system, according to the manufacturer’s instructions. Immunoblotting was performed using antibodies for heme oxygenase-1 (HMOX1) (catalog no. 13248, Abcam; 1:250) and thrombospondin-1 (THBS1) (catalog no. 1823, Abcam; 1:1,000) with α-mouse secondary (catalog no. 925-32210, LICOR; 1:15,000). β-Actin was probed using a 1°-horseradish peroxidase (HRP) antibody (MA5-15739, Thermo Scientific; 1:1,000) and developed using ECL Plus (Thermo Scientific). Detection of blots was performed using a LICOR Odyssey LC system and quantified using Image Studio (v.4.0, LICOR).

Shearer J.J., Wold E.A., Umbaugh C.S., Lichti C.F., Nilsson C.L, & Figueiredo M.L. (2015). Inorganic Arsenic–Related Changes in the Stromal Tumor Microenvironment in a Prostate Cancer Cell–Conditioned Media Model. Environmental Health Perspectives, 124(7), 1009-1015.

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Embryonic Western Blot Analysis

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Western blot analysis was performed as described previously (Dickinson and Sive, 2009 (link)). Briefly, embryos were flash frozen in liquid nitrogen and stored or immediately immersed in lysis buffer with protease inhibitors (Sigma Aldrich, S8820). Samples were centrifuged and the protein containing aqueous fraction was obtained by piercing the tube with an 18G needle and extracting said fraction with a syringe. Primary antibodies used were: β-Actin (Cell Signaling, 4970S, 1:1000), β-Actin, (Sigma Aldrich, A5441, 1:5000), histone H3 (Abcam, ab24834, 1:5000), and tri-methyl-histone-H3 (Cell Signaling, 9751S, 1:1000). Secondary antibodies included goat anti-rabbit IgG (Cell Signaling, 5151S) or goat anti-mouse IgG IRDye680 (LICOR, 926–68070) diluted (1:6667). Visualization and quantification was done using the Odyssey CLx Infrared Imaging System and requisite software (Scanning: Image Studio v.4.0, Quantification: Image Studio Lite v.4.0, LI-COR Biosciences).

Wahl S.E., Kennedy A.E., Wyatt B.H., Moore A.D., Pridgen D.E., Cherry A.M., Mavila C.B, & Dickinson A.J. (2015). The role of folate metabolism in orofacial development and clefting. Developmental biology, 405(1), 108-122.

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Protein Expression Analysis by Western Blot

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Cell pellets (5 × 10⁶ cells) were lysed in RIPA buffer (Thermo Fisher Scientific) supplemented with protease inhibitors (Sigma-Aldrich) for 15 min on ice. Total protein content was determined using the Qubit protein assay kit (Thermo Fisher Scientific) following the manufacturer´s protocol. Protein lysates (40 µg/lane) were separated by SDS-PAGE and transferred to polyvinylidene fluoride membranes (BioRad). Membranes were blocked with Intercept (TBS) blocking buffer (LI-COR) for 1 h at RT and incubated with primary antibodies mCherry (E5DF8, #43590) and β-Tubulin (9F3, #2128) (Cell Signalling Technologies) diluted 1:1000 in blocking buffer at 4 °C overnight. Membranes were washed three times with 0.1% TBS-Tween and incubated for 1 h at RT with IRDye 800CW goat anti-rabbit IgG cat# 926–32211 (LI-COR) diluted 1:20,000 in 0.1% TBS-Tween. Following washes, membranes were visualised with the Odyssey CLx imaging system and processed in Image Studio (v4.0, LI-COR).

Munson M.J., O’Driscoll G., Silva A.M., Lázaro-Ibáñez E., Gallud A., Wilson J.T., Collén A., Esbjörner E.K, & Sabirsh A. (2021). A high-throughput Galectin-9 imaging assay for quantifying nanoparticle uptake, endosomal escape and functional RNA delivery. Communications Biology, 4, 211.

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Immunoblotting Analysis of Signaling Proteins

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For immunoblotting analysis, protein extraction from cultured COS7 cells was conducted according to previously described methods [24 (link)]. Whole-cell lysates were separated using SDS/10% polyacrylamide gels and electrophoretically transferred to a polyvinylidene fluoride membrane (Bio Trace NT Nitrocellulose Transfer Membrane, Pall Corporation, New York, NY, USA) using standard procedures. Membranes were blocked with 3% weight/volume skim milk in TBS and probed with primary antibodies against FGFR2 (1:2000 dilution, Cell Signaling Technologies, Danvers, MA, USA), phospho and total 44/42 MAPK (1:1000 dilution; Cell Signaling Technologies, Danvers, MA, USA), pFRS2 (1:500 dilution, Cell Signaling Technologies, Danvers, USA), and FRS2 (1:2000 dilution, Proteintech, Rosemont, IL, USA) followed by incubation at 4 °C for 12 h. After three washes in 0.1% Tween-TBS, the membranes were incubated with secondary antibody at 37 °C for 1 h. Peroxidase-conjugated goat anti-mouse or goat anti-rabbit antibodies (Promega, Madison, WI, USA) were used as the secondary antibodies. Immunoreactive protein bands were visualized using ECL chemifluorescent reagent (GE Healthcare, Chicago, CA, USA) and imaged with Image Studio v4.0 (LI-COR, Lincoln, USA) in the LI-COR Odyssey FC infrared imager.

Dixit G., Pappas B.A., Bhardwaj G., Schanz W, & Maretzky T. (2023). Functional Distinctions of Endometrial Cancer-Associated Mutations in the Fibroblast Growth Factor Receptor 2 Gene. Cells, 12(18), 2227.

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Lipid Nanoparticle Characterization

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2 µL solution containing LNPcor or corresponding control samples was spotted to Nitrocellulose membranes (#LC2001, Thermo Fisher Scientific). Membranes were blocked with Intercept (TBS) blocking buffer (LI-COR) for 30 min at RT and incubated with primary antibodies Cy5 (#ab52061, Abcam), PEG (#ab51257, Abcam), Apo AII (#ab92478, Abcam), Apo CII (#ab230447, Abcam), Apo CIII (#ab76305, Abcam) and ApoE (#ab183597, Abcam) where appropriate, diluted 1:1000 in blocking buffer at for 30 min at RT. Membranes were washed three times with 0.1% TBS-Tween and incubated for 30 min at RT with IRDye 680RD Goat anti-Mouse IgG (#926-68070, LI-COR Biotechnology) and 800CW goat anti-rabbit IgG (#926-32211, LI-COR Biotechnology) diluted 1:2000 in 0.1% TBS-Tween. Following three washes, membranes were visualized with the Odyssey CLx imaging system and processed in Image Studio (v4.0, LI-COR).

Liu K., Nilsson R., Lázaro-Ibáñez E., Duàn H., Miliotis T., Strimfors M., Lerche M., Salgado Ribeiro A.R., Ulander J., Lindén D., Salvati A, & Sabirsh A. (2023). Multiomics analysis of naturally efficacious lipid nanoparticle coronas reveals high-density lipoprotein is necessary for their function. Nature Communications, 14, 4007.

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Protein Extraction and Western Blot Analysis

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Cells and tissues were lysed in lysis buffer (0.1 M Tris‐HCl pH 6.8, 4% SDS, 20% glycerol, sodium pyrophosphate, 1 mM Na3VO4, NaF) and applied to 10% SDS‐PAGE gels and probed with antibodies. Blots were visualized by using the LI‐COR Odyssey System (Lincoln, NE). Quantitative determination of band intensity involved using Image Studio v4.0 (LI‐COR). Antibodies are listed in Supporting Table S1.

Zhang J., Li Y., Liu Q., Huang Y., Li R., Wu T., Zhang Z., Zhou J., Huang H., Tang Q., Huang C., Zhao Y., Zhang G., Jiang W., Mo L., Zhang J., Xie W, & He J. (2020). Sirt6 Alleviated Liver Fibrosis by Deacetylating Conserved Lysine 54 on Smad2 in Hepatic Stellate Cells. Hepatology (Baltimore, Md.), 73(3), 1140-1157.

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Quantitative Extracellular Vesicle Protein Analysis

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Extracellular vesicle pellets were resuspended in 100 μl of PBS. For immunoblotting, a 10 μl aliquot was mixed with an equal volume of 2X radioimmunoprecipitation assay buffer and incubated on ice for 30 min. SDS-PAGE sample buffer was added and samples were incubated at 95 °C for 5 min.
Proteins were resolved by 4-15% SDS-PAGE and transferred to a Immobilon-FL PVDF membrane (MilliporeSigma, catalog no. IPFL20200). The membrane was blocked with a solution of 5% BSA in TBST (50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.1% Tween 20) for 1 h, then incubated overnight at 4 °C with primary antibodies diluted in a solution of 1% BSA in TBST. Primary antibodies were as follows: anti-TSG101 (Abcam, Cambridge, MA; catalog no. ab83, 1:1000), anti-calreticulin (Abcam, catalog no. ab2907, 1:1000), and anti-HSP70 (Enzo Life Sciences, Inc., Farmingdale, NY, cat no. ADI-SPA-810-D, 1:1000). Membranes were washed with TBST, then incubated with a solution of 1% BSA in TBST containing a 1:15,000 dilution of IR Dye 680RD-conjugated goat anti-mouse IgG (LI-COR Biosciences, Lincoln, NE, catalog no. 925-68070). Immune complexes were imaged using a LI-COR Odyssey system and quantified using Image Studio v4.0 (LI-COR Biosciences).

Li Z., Jella K.K., Jaafar L., Moreno C.S., & Dynan W.S. (2020). Characterization of exosome release and extracellular vesicle-associated miRNAs for human bronchial epithelial cells irradiated with high charge and energy ions.

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Neuroretinal Protein Expression Analysis

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Neuroretinal extracts were prepared from 3 normal, 3 rcd1 and 2 xlpra2 retinas in a buffer containing 0.23 M sucrose, 2 mM EDTA, 5 mM Tris HCl, pH 7.4, 1% Tx100 and a protease and phosphatase inhibitor cocktail (Halt, Thermo Scientific, Waltham, MA). The retinas were first homogenized by vortexing with 1.5 mm zirconium beads (Benchmark Scientific Inc., Sayreville, NJ) and then sonicated. The samples were then centrifuged and total protein concentration in the supernatant measured by BCA assay. 50 µgs of total protein from each sample was resolved on an 8–16% Tris Glycine gel (Invitrogen, Carlsbad, CA), transferred to a nitrocellulose membrane (iBLOT, Invitrogen) and immunoblotted using antibodies listed in Supplementary Table 6. After incubation with infrared dye-tagged secondary antibodies, protein bands were visualized on a digital imaging system (Odyssey Fc, Licor, Lincoln, NE). Quantification was performed using the Licor Image Studio v4.0 software using β-actin bands for normalization. Statistical significance was calculated for rcd1 samples only by unpaired homoscedastic t-test using a two-tailed distribution.

Sudharsan R., Beiting D.P., Aguirre G.D, & Beltran W.A. (2017). Involvement of Innate Immune System in Late Stages of Inherited Photoreceptor Degeneration. Scientific Reports, 7, 17897.

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