Hydrogel composition to enhance fluorescence

• US 8,383,415 B2
• Filed: 10/13/2005
• Issued: 02/26/2013
• Est. Priority Date: 10/13/2005
• Status: Expired due to Fees

• Current Assignee: DSO National Laboratories
• Original Assignee: DSO National Laboratories
• Inventors: Ayi, Teck Choon, Tong, Mei-Mei Jill, Lee, Vee Sin Peter
• Primary Examiner(s): Xu, Robert
• Application Number:
  US12/083,593
• Publication Number:
  US 20090130773A1
• Time in Patent Office: 2,693 Days
• Field of Search: None
• US Class Current: 436/172
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 2563/107 fluorescence
• CPC Class Codes: C12Q 2563/149 Particles, e.g. beads
• CPC Class Codes: C12Q 2563/179 the label being a nucleic acid
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 33/544 the carrier being organic
• CPC Class Codes: G01N 33/582 with fluorescent label

Abstract

A composition comprising a hydrogel particle and a fluorophore; wherein said composition produces an enhanced fluorescent signal when excited by an energy source capable of exciting the fluorophore.

Claims

1. A composition comprising:

a hydrogel material comprising hydrogel particles crosslinked by enzyme-degradable crosslinkers made from a protein, polysaccharide, peptide or polypeptide and a fluorophore incorporated into the crosslinkages of the hydrogel material, wherein said composition produces an enhanced fluorescent signal when excited by an energy source capable of exciting the fluorophore, wherein the enhanced fluorescent signal generated by the composition of the hydrogel particle and a fluorophore said intensity is greater than the intensity of a fluorescent signal generated by a composition comprising the fluorophore in the absence of the hydrogel particle.

2. The composition of claim 1 wherein the hydrogel particles are formed from poly N-isopropyl-acrylamide(pNIPAM).

3. The composition of claim 2 wherein the pNIPAM is polymerised with hydroxyethyl acrylate to form pNIPAM-co-hydroxyethyl acrylate (pNIPAM-HEA).

4. The composition of claim 2 wherein the pNIPAM is polymerised with acrylic acid to form pNIPAM-co-acrylic acid (pNIPAM-AA).

5. The composition of claim 2 wherein the pNIPAM is polymerised with allylamine to form pNIPAM-co-allylamine acid (pNIPAM-Allylamine).

6. A composition of claim 2 wherein the pNIPAM is polymerised with at least an unsaturated double bond monomer to form a copolymer of pNIPAM which includes functional groups for chemical reactions.

7. A composition of claim 6 wherein the functional group is selected from the group comprising:

an amine, a sulfurhydryl, a carboxyl, a hydroxyl, and an aromatic ring structure.

8. The composition of claim 1 wherein the hydrogel particle is a hydrogel nanoparticle.

9. The composition of claim 1 further comprising a liquid media and said composition is a colloidal suspension.

10. The composition of claim 9 wherein the liquid media is chosen from the list comprising:

water, phosphate buffer saline (PBS), 1×

Tris-EDTA.

11. The composition according to claim 1 wherein the crosslinks are environmentally responsive.

12. The composition according to claim 11 wherein the environmentally responsive crosslinks respond to an environmental factor by contracting, expanding, or degrading.

13. The composition according to claim 12 wherein the environmental factor is selected from the group consisting of temperature, chemical, enzyme, microbial action, enzymatic action, and mixture thereof.

14. The composition according to claim 13 further comprising a growth media for bacterial growth.

15. The composition according to claim 14 wherein the microbial action degrades the hydrogel material, thereby liberating the hydrogel particles and fluorophores.

16. The composition according to claim 1 wherein the crosslinks are bovine serum albumen.

17. The composition according to claim 1 wherein the fluorophore is an extrinsic fluorophore.

18. The composition according to claim 17 wherein the extrinsic fluorophore is selected from the list comprising:

Alexa Fluor®

350, Dansyl Chloride (DNS-CI), 5-(iodoacetamida)fluoroscein (5-IAF);

fluorescein 5-isothiocyanate (FITC), tetramethylrhodamine 5-(and 6-)isothiocyanate (TRITC), 6-acryloyl-2-dimethylaminonaphthalene (acrylodan), 7-nitrobenzo-2-oxa-1,3,-diazol-4-yl chloride (NBD-CI), ethidium bromide, Lucifer Yellow, 5-carboxyrhodamine 6G hydrochloride, Lissamine rhodamine B sulfonyl chloride, Texas Red™

sulfonyl chloride, BODIPY™

, naphthalamine sulfonic acids, 1-anilinonaphthalene-8-sulfonic acid (ANS),6-(p-toluidinyl)naphthalene-2-sulfonic acid (TNS), Anthroyl fatty acid, DPH, Parinaric acid, TMA-DPH, Fluorenyl fatty acid, Fluorescein-phosphatidylethanolamine, Texas red-phosphatidylethanolamine, Pyrenyl-phophatidylcholine, Fluorenyl-phosphotidylcholine, Merocyanine 540, 1-(3-sulfonatopropyl)-4[-β

-[2-[(di-n-butylamino)-6naphthyl]vinyl]pyridinium betaine (Naphtyl Styryl), 3,3′

dipropylthiadicarbocyanine (diS-C₃-(5)), 4-(p-dipentyl aminostyryl)-1-methylpyridinium (di-5-ASP), Cy-3 Iodo Acetamide, Cy-5-N-Hydroxysuccinimide, Cy-7-Isothiocyanate, rhodamine 800, IR-125, Thiazole Orange, Azure B, Nile Blue, Al Phthalocyanine, Oxaxine 1,4′

,6-diamidino-2-phenylindole (DAPI), Hoechst 33342, TOTO, acridine orange, ethidium homodimer, N(ethoxycarbonylmethyl)-6-methoxyquinolinium (MQAE), Fura-2, Calcium Green, Carboxy SNARF-6, BAPTA, coumarin, phytofluors, Coronene, metal-ligand complexes, quantum dots, green fluorescent protein, yellow fluorescent protein, red fluorescent protein, mutants and derivates thereof.

19. The composition according to claim 18 wherein the extrinsic fluorophore is selected from the list comprising:

ethidium bromide, Sybr Green 1 and fluorescein.

20. The composition according to claim 1 wherein the fluorophore is an intrinsic fluorophore.

21. The composition according to claim 20 wherein the intrinsic fluorophore is selected from the list comprising:

tyrosine, tryptophan, phenylalanine and proteins and peptides with these amino acids, NADH, NADPH, FAD, vitamins, enzyme-cofactors, lanthanide, purines, pyrimidines, lipids, fatty acids, nucleic acids, amino acids, carbohydrates, steroids, flavins, oligonucleotides, aromatic carbon ring structures, nucleotides, nucleosides, proteins, peptides, DNA, RNA, sugars, purines, pyrimidines, formycin, phytochrome, phytofluor, antibodies, phycobiliprotein, and mixture thereof.

22. The composition according to claim 21 wherein the intrinsic fluorophore is a protein.

23. The composition according to claim 1 wherein the fluorophore is conjugated to the hydrogel particle.

24. The composition according to claim 1 wherein the fluorophore is carried inside the hydrogel particle.

25. The composition according to claim 1 wherein the fluorophore is carried inside the hydrogel material.