Amine-PEG3-Biotin

Amine-PEG3-Biotin is a signal amplification label containing a biotin group and a terminal primary amine group^[1]. In Vitro:Guidelines (The following is our recommended protocol. This protocol is only a guide and should be modified to suit your specific needs).
1 Preparation of dye working solution^[1]:
1) 1 mL of Amine-PEG3-Biotin working solution at a concentration of 5 mg/mL is used. If starting from a stock solution, calculate the volume of stock solution required and dilute to the desired concentration with 10 mM PBS (pH7.4). For example, if the stock solution concentration is 10 mg/mL, take 0.5 mL of stock solution and add 0.5 mL of 10 mM PBS (pH7.4) to obtain a 5 mg/mL working solution.
2 Specific modification steps:
1) Disperse 5 mg of carboxyl-functionalized SiO₂ nanospheres in 50 mL of deionized water under ultrasound assistance and sonicate for 30 min. Then, the dispersion is added to a 250 mL round-bottom flask and magnetically stirred.
2) 10 mL of 3 mM AgNO₃ is added to the round-bottom flask and the dispersion is stirred for 1 h.
3) 5 mL of 6 mM sodium citrate is quickly added to the flask and boiled for 30 min.
4) The obtained suspension is centrifuged at 8000 r/min for 20 min, the precipitate is collected, washed three times with deionized water, and vacuum dried. 5) 1 mg of the obtained Ag@SiO₂ nanocomposite is dispersed in 0.5 mL of 10 mM PBS (pH7.4) under ultrasound.
6) Add 0.5 mL of a mixed solution containing 50 mM EDC and 100 mM NHS to the Ag@SiO₂ dispersion and incubate with shaking for 1 h.
7) Add 1 mL of 5 mg/mL Amine-PEG3-Biotin to the dispersion and react for 1 h.
8) Centrifuge the mixture and incubate the precipitate with 1 mL of 0.1 mM ethanolamine for 1 h to remove the carboxyl residues on the SiO₂ surface.
9) Finally, collect the biotin-functionalized Ag@SiO₂ nanocomposites by centrifugation at 8000 r/min and vacuum dry the resulting precipitate.