Hello,
I am a student from İstanbul Technical University.
I would like to ask that which parameters should be considered by choosing method for conjugation between antibody and gold nanoparticle? Which details should I pay attention?
Does method changes based on antibody?
Thanks in advance.
1st. what is your application.
2nd. what size of beads you want to use (depend on decision 1).
How many antibodies per beads.
There are commercial kits that are very easy to use, but also the simple methods are easy (more time consuming), but after you preper your beads you can save them at room temp. before conjugation.
size of beads can be mesured by absorbance.
good luck
KLH (keyhole cap hemocyanin) is the most commonly used carrier protein, which has higher immunogenicity compared with other proteins. KLH is commonly used in vertebrate research.
BSA (Bovine serum albumin) is a commonly used weak antigenic compound carrier protein because it contains a large number of binding lysine residues.
HSA (Human serum albumin)
OVA (Ovalbumin) is a protein isolated from eggs. It is often used as a control carrier protein to verify whether the antibodies produced by KLH/BSA are specific to target peptide and carrier proteins.
I presume you are making gold-antibody conjugates for lateral flow test. You need to address the following points:
1) use 40nm spherical gold particles.
2) different antibodies behave differently. Do trial conjugation using 3 different pH. I suggest pH 7.5, 8.5 and 9.5. borate buffer at 25mM is the best. block with BSA
3) avoid high concentration of NaCl. Salt cause collapse of the gold particles.
4) use a ratio of 2ug IgG antibody per 1 optical density of gold. With this ratio, in most cases, you will not need to wash conjugate by centrifugarion to remove unbound antibody.
Falk Fish, could you please provide articles that contain established protocols for analytical approaches to confirm gold nanoparticle antibody conjugation? and do you have the aforementioned books in PDF format if so could you please send me those?
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