Biotinylated-BSA (1 μg/μl, Thermo) was flowed for 30 min through the device, binding the BSA to the epoxy surface. On top of the biotinylated-BSA, 0.5 μg/μl of Neutravidin (Pierce, Rockford, IL) was added for 30 min. The button valve was then closed, and biotinylated-PEG (1 μg/μl, Nanocs) was flowed over for 30 min, thus passivating the rest of the flow layer. Following passivation, the button valve was released and a flow of 0.2 μg/μl penta-His biotinylated (Qiagen, Venlo, Netherlands) or 0.01 µg/µl anti-Myc biotinylated antibodies (Cell Signaling, Danvers, MA, USA) were applied. The antibodies bound to the exposed Neutravidin, specifically to the area under the button, creating an array of anti-His- or anti-Myc tag. PBS buffer was used for washing between each surface chemistry step.
Biotinylated peg
Manufactured by Nanocs
Biotinylated-PEG is a versatile laboratory reagent designed for various bioconjugation applications. It consists of a polyethylene glycol (PEG) backbone with biotin moieties attached, which can be utilized to bind target molecules or surfaces. The biotinylation property allows for efficient and specific interactions with streptavidin or avidin-based systems.
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Lab products found in correlation
2 protocols using biotinylated peg
1
Fabrication of Protein Microarray using Microfluidics
2
Protein Capture Using Biotinylated Antibodies
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To bind the expressed protein, we first cover the epoxy slides with biotinylated BSA (1 µg/µl; Thermo Scientific). Streptavidin (0.5 µg/µl Neutravidin; Pierce) was then introduced to interact with the biotinylated BSA layer. Next, a designated set of pneomatic valves, also known as “button valves” (Noach-Hirsh et al., 2015 (link)), were closed and a layer of biotinylated PEG (1 µg/µl; Nanocs) was used to block the protein chamber periphery, leaving the center itself exposed to avidin to which antibodies and eventually target proteins can be attached. Then, the button valves were opened and anti-GFP biotinylated antibodies (0.2 µg/µl; Abcam; #ab6658) were flowed through the device to interact with the exposed avidin at the protein chamber center. This creates an array of protein chamber with anti GFP antibodies to which EGFP-tagged E2F8 IVT product can bind. PBS buffer or PBS buffer with 1% BSA was used to the wash between steps.
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