I am working to functionalize AuNPs with oligonucleotides. the problem is how I can show the conjugation between oligonucleotides and AuNPs, by using which technique? In theoritically, thiolated oligonucleotides are surrounded AuNP surface. But, which technique can help me to prove that they are on the surface of AuNP?
Thanks for answers :)
the classical approach would be to separate the nanoparticles and determine concentration of oligonucleotides in solution used for conjugation.
If you have purified the free oligonucleotides from your nanoparticles, you can accomplish this by mixing your nanoparticle solution with KCN (100 uM) until the color disappears. Then, run the mixture through a gel filtration column (NAP-5 or NAP-10) and collect the fractions. Determine the total concentration of DNA in each fraction by UV-Vis. The total amount of particles divided by total amount of DNA will give you the number of oligonucleotides/NP.
Melike,
you can use several techniques. Check inh literature the position of surface plasmon after conjugation. It should be red shifted and slightly wider. Second, you can do DLS/Zeta check. In the case of oligonucleotide coupling you observe negative charge compared to positively charged Au-CTAB (If it's CTAB). Moreover DLS size should be larger in the case of conjugation compared to initial particles. In the end you can do gel electrophoresis and stain it with ethidium bromide, for example.
Regards,
Kostia
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