Assays for measuring binding kinetics and binding capacity of acceptors for lipophilic or amphiphilic molecules

Abstract

Aspects of the invention relate to methods for measuring the binding constant of a lipophilic or amphiphilic molecule acceptor for a lipophilic or amphiphilic molecule. Methods involve rapid, cell-free competition assays including a labeled lipophilic or amphiphilic molecule and nanoparticle.

What is claimed is: 
     
         1 .- 9 . (canceled) 
     
     
         10 . A kit for measuring an equilibrium constant of an acceptor for a lipophilic or amphiphilic molecule, comprising:
 (a) the lipophilic or amphiphilic molecule having a detectable signal; and   (b) a structure, the structure comprising a nanostructure core and a lipid layer surrounding and attached to the nanostructure core, wherein the structure quenches the signal of the lipophilic or amphiphilic molecule when the structure and the lipophilic or amphiphilic molecule are proximate.   
     
     
         11 . The kit of  claim 10 , further comprising instructions for use of the kit for measuring the equilibrium constant of the acceptor for the lipophilic or amphiphilic molecule. 
     
     
         12 . A system for measuring an equilibrium constant of an acceptor for a lipophilic or amphiphilic molecule, comprising:
 (a) a sample, the sample comprising:
 (i) a lipophilic or amphiphilic molecule, wherein the molecule can provide a detectable signal; 
 (ii) a structure, the structure comprising a nanostructure core and a lipid layer surrounding and attached to the nanostructure core, wherein the structure quenches the signal of the lipophilic or amphiphilic molecule when the structure and the lipophilic or amphiphilic molecule are proximate; 
 (iii) an acceptor; and 
   (b) a detector for measuring the signal.   
     
     
         13 . The system of  claim 12 , further comprising a device configured to calculate the equilibrium constant of the acceptor for the lipophilic or amphiphilic molecule based on the detected signal. 
     
     
         14 . The system of  claim 12 , further comprising a radiation source configured to induce the signal. 
     
     
         15 .- 32 . (canceled) 
     
     
         33 . The kit of  claim 10 , wherein the equilibrium constant is measured in a cell-free assay. 
     
     
         34 . The kit of  claim 10 , wherein the lipophilic or amphiphilic molecule comprises a steroid, a lipopolysaccharide, a cholestane, cholane, pregnane, androstane, estrane or a derivative or analog thereof. 
     
     
         35 . The kit of  claim 10 , wherein the lipophilic or amphiphilic molecule comprises or consists of BODIPY-cholesterol. 
     
     
         36 . The kit of  claim 10 , wherein the signal is fluorescence. 
     
     
         37 . The kit of  claim 10 , wherein the nanostructure core is an inorganic material. 
     
     
         38 . The kit of  claim 10 , wherein the nanostructure core is gold. 
     
     
         39 . The kit of  claim 10 , wherein the structure further comprises an apolipoprotein bound to at least the outer surface of the lipid layer. 
     
     
         40 . The kit of  claim 10 , wherein the acceptor is a high-density lipoprotein (HDL). 
     
     
         41 . The system of  claim 12 , wherein the sample is a biological sample and wherein the biological sample is not a cell. 
     
     
         42 . The system of  claim 12 , wherein the lipophilic or amphiphilic molecule comprises a steroid, a lipopolysaccharide, a cholestane, cholane, pregnane, androstane, estrange, or BODIPY-cholesterol. 
     
     
         43 . The system of  claim 12 , wherein the signal is detected by positron emission tomography, SPECT medical imaging, chemiluminescence, electron-spin resonance, ultraviolet/visible absorbance spectroscopy, mass spectrometry, nuclear magnetic resonance, magnetic resonance, flow cytometry, autoradiography, scintillation counting, phosphoimaging, or an electrochemical method. 
     
     
         44 . The system of  claim 12 , wherein the signal is fluorescence. 
     
     
         45 . The system of  claim 12 , wherein the nanostructure core is gold. 
     
     
         46 . The system of  claim 12 , wherein the structure further comprises apolipoprotein bound to at least the outer surface of the lipid layer. 
     
     
         47 . The system of  claim 12 , wherein the acceptor is a high-density lipoprotein (HDL).