Protein concentration was measured using a Direct Detect infrared spectrometer (Merck). Twenty micrograms of proteins was separated on a NuPAGE 4%–12% Bis-Tris gel (Thermo Fisher Scientific). Proteins were transferred with an iBLot2 device (Invitrogen) on a nitrocellulose membrane and blocked for 1 h in 1× Licor TBS blocking buffer (Licor). Primary antibodies were incubated overnight at 4°C. Licor secondary antibodies were incubated for 45 min at room temperature, and images were acquired with an Odyssey CLx scanner (Licor) using secondary antibodies conjugated to infrared dyes from LiCor. The following primary antibodies were used: anti-CG13741, anti-Nxf3, anti-Tubulin (ab18251), anti-GFP (ab13790), anti-Flag (Sigma, F1804), and anti-HA (Cell Signaling, 3724S).
Licor tbs blocking buffer
Manufactured by LI COR
Sourced in United States
LI-COR TBS Blocking Buffer is a pre-formulated blocking solution for Western blotting applications. It is designed to minimize non-specific binding of antibodies to the membrane, improving signal-to-noise ratio and reducing background.
Automatically generated - may contain errors
Lab products found in correlation
Direct detect infrared spectrometer, by Merck Group (4 mentions)
Odyssey clx scanner, by LI COR (4 mentions)
Nupage 4 12 bis tris gel, by Thermo Fisher Scientific (4 mentions)
Anti flag, by Merck Group (2 mentions)
Iblot2 device, by Thermo Fisher Scientific (2 mentions)
Immun blot low fluorescent pvdf membrane, by Bio-Rad (2 mentions)
Anti tubulin, by Merck Group (1 mentions)
Anti gfp, by Merck Group (1 mentions)
Anti ha, by Cell Signaling Technology (1 mentions)
Odyssey imaging system, by LI COR (1 mentions)
Bis tris sds page gel, by Thermo Fisher Scientific (1 mentions)
Infrared tagged secondary antibodies, by LI COR (1 mentions)
Iblot 2 dry blotting system, by Thermo Fisher Scientific (1 mentions)
Irdye 800cw donkey anti rabbit igg, by LI COR (1 mentions)
Odyssey scanner, by LI COR (1 mentions)
Ripa buffer, by Santa Cruz Biotechnology (1 mentions)
Irdye 800cw donkey anti mouse igg, by LI COR (1 mentions)
Irdye 680rd donkey anti rabbit igg, by LI COR (1 mentions)
Irdye 800cw goat anti rat igg, by LI COR (1 mentions)
6 protocols using licor tbs blocking buffer
1
Protein Quantification and Western Blot
2
Western Blot Protein Separation and Detection
3
Western Blot Analysis of Extracellular Vesicles
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SF-sEVs, PC3 sEVs, and PC3 cells were lysed in RIPA buffer (Santa Cruz Biotechnology, Santa Cruz, CA) supplemented with protease inhibitors, separated by SDS-PAGE under reducing conditions, and blotted with iBlot2 dry blotting system (ThermoFisher Scientific). LI-COR TBS blocking buffer (LI-COR Biosciences, Lincoln, NE, USA) was used for blocking and antibody incubation. The proteins were analyzed using 1.3 µg/ml anti-TSG101 antibody, 0.5 µg/ml anti-CD9 antibody, 0.5 µg/ml anti-CD81 antibody, 0.5 µg/ml anti-CD63 antibody and 0.1 µg/ml anti-calnexin antibody, which were detected using 50 ng/ml donkey anti-mouse IgG IRDye 680LT or 75 ng/ml donkey anti-rabbit IgG IRDye 800CW as secondary antibodies and analyzed using Odyssey scanner from LI-COR. All information for the antibodies are listed in Supplementary Table 1 .
4
Immuno-blotting of protein samples
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Protein concentration was measured using a Direct Detect Infrared Spectrometer (Merck). 20 µg of proteins were separated on a NuPAGE 4–12% Bis-Tris gel (Thermo Fisher Scientific). Proteins were transferred with an iBLot2 device (Invitrogen) on a nitrocellulose membrane and blocked for 1 hr in 1x Licor TBS Blocking Buffer (Licor). Primary antibodies were incubated over night at 4°C. Licor secondary antibodies were incubated for 45 min at room temperature (RT) and images acquired with an Odyssey CLx scanner (Licor). The following antibodies were used: anti-HA (ab9110), anti-FLAG (Sigma #F1804), anti-GFP (ab13970), anti-Piwi (described in Brennecke et al., 2007 (link)), anti-Nxt1 (Herold et al., 2001 (link)), anti-Histone H3 (ab10799), anti-Tubulin (ab18251), mouse anti-Panx (Sienski et al., 2015 (link)), IRDye 680RD Donkey anti-Rabbit IgG (H + L) (Licor), IRDye 800CW Donkey anti-Mouse IgG (H + L) (Licor), IRDye 800CW Goat anti-Rat IgG (H + L) (Licor).
5
Protein Quantification and Western Blotting
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Protein concentration was measured using a Direct Detect Infrared Spectrometer (Merck). 20 µg of proteins were separated on a NuPAGE 4–12% Bis-Tris gel (Thermo Fisher Scientific). Proteins were transferred for 2 hr at 100 V, 400 mA, 100 W on an Immun-Blot Low Fluorescent PVDF Membrane (BioRad) and blocked for 1 hr in 1× LI-COR TBS Blocking Buffer (LI-COR). Primary antibodies were incubated overnight at 4°C. LI-COR secondary antibodies were incubated for 45 min at room temperature (RT) and images acquired with an Odyssey CLx scanner (LI-COR).
6
Quantitative Protein Analysis by Western Blot
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Protein concentration was measured using a Direct Detect Infrared Spectrometer (Merck). 20µg of proteins were separated on a NuPAGE 4-12% Bis-Tris gel (Thermo Fisher Scientific). Proteins were transferred for 2h at 100V, 400mA, 100W on an Immun-Blot Low Fluorescent PVDF Membrane (BioRad) and blocked for 1h in 1x Licor TBS Blocking Buffer (Licor). Primary antibodies were incubated over night at 4°C. Licor secondary antibodies were incubated for 45min at room temperature (RT) and images acquired with an Odyssey CLx scanner (Licor).
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