Antibody-nanoparticle conjugates and methods for making and using such conjugates
Disclosed herein are antibody-nanoparticle conjugates that include two or more nanoparticles (such as gold, palladium, platinum, silver, copper, nickel, cobalt, iridium, or an alloy of two or more thereof) directly linked to an antibody or fragment thereof through a metal-thiol bond. Methods of making the antibody-nanoparticle conjugates disclosed herein include reacting an arylphosphine-nanoparticle composite with a reduced antibody to produce an antibody-nanoparticle conjugate. Also disclosed herein are methods for detecting a target molecule in a sample that include using an antibody-nanoparticle conjugate (such as the antibody-nanoparticle conjugates described herein) and kits for detecting target molecules utilizing the methods disclosed herein.
1. A kit for detecting a target molecule in a sample, comprising one or more containers containing a first antibody-nanoparticle conjugate, the first antibody-nanoparticle conjugate comprising three or more metal nanoparticles directly linked to a single antibody through three or more metal-thiol bonds, wherein the metal-thiol bonds are between thiol from cysteine residues of the antibody and the three or more metal nanoparticle without an intervening linker.
2. The kit of claim 1 , further comprising a second antibody conjugated to one or more enzyme molecules, wherein the first antibody specifically binds to the second antibody.
3. The kit of claim 1 , wherein the three or more nanoparticles comprise gold, palladium, platinum, silver, copper, nickel, cobalt, iridium, or an alloy of two or more thereof.
4. The kit of claim 1 , wherein the three or more nanoparticles are about 5 nm or less in diameter.
5. The kit of claim 4 , wherein the three or more nanoparticles are about 0.5-5 nm in diameter.
6. The kit of claim 1 , wherein the first antibody is conjugated to three to seven gold nanoparticles.
7. The kit of claim 2 , wherein the second antibody comprises a goat-anti-rabbit antibody.
8. The kit of claim 7 , wherein the first antibody is a rabbit anti-goat antibody.
9. The kit of claim 2 , wherein the one or more enzyme molecules comprise an alkaline phosphatase, an acid phosphatase, a β-galactosidase, a β-glucosidase, a β-lactamase, or an esterase.
10. The kit of claim 9 , wherein the one or more enzyme molecules is alkaline phosphatase.
11. The kit of claim 10 , wherein the second antibody is conjugated to three alkaline phosphate molecules.
12. The kit of claim 2 , wherein the kit further comprises one or more containers comprising a substrate for the enzyme and a metal ion.
13. The kit of claim 12 , wherein the metal ion comprises silver ion, gold ion, copper ion, nickel ion, platinum ion, palladium ion, cobalt ion, or iridium ion.
14. The kit of claim 12 , wherein the substrate for the enzyme comprises 5-bromo-4-chloro-3-indolyl phosphate, ascorbic acid phosphate, or a hydroquinone phosphate derivative.
15. The kit of claim 1 , further comprising a gold halide.
16. The kit of claim 15 , wherein the gold halide comprises gold chloride.
17. The kit of claim 1 , further comprising a silver salt.
18. The kit of claim 17 , wherein the silver salt comprises silver nitrate, silver oxide, or silver chloride.
19. The kit of claim 1 , further comprising a reducing agent.
20. The kit of claim 19 , wherein the reducing agent comprises sodium thiosulfate.
21. The kit of claim 1 , further comprising a third antibody that specifically binds a target molecule, wherein the second antibody specifically binds to the third antibody.