Systems and methods for high throughput analysis of conformation in biological entities

Abstract

Methods, devices, and systems are disclosed for performing high throughput analysis of conformational change in biological molecules or other biological entities using surface-selective nonlinear optical detection techniques.

1 . A method for determining interactions between target molecules and test compounds, comprising:
 a) contacting at least one target molecule with at least one test compound, wherein the at least one target molecule is labeled with a nonlinear-active label and is tethered to at least one discrete region on an upper surface of a planar substrate;   b) illuminating the at least one target molecule with one or more excitation light beams using total internal reflection of the one or more excitation light beams from the upper surface of the planar substrate, wherein the planar substrate comprises an array of prisms integrated with a lower surface, wherein the one or more excitation light beams are directed by an entrance prism to a single discrete region that is adjacent to but not directly above the entrance prism, wherein the one or more reflected excitation light beams and a nonlinear optical signal generated by the at least one labeled target molecule tethered to the single discrete region are collected by an exit prism that is also adjacent to but not directly underneath the single discrete region, and wherein the entrance prism and the exit prism for the single discrete region are different, non-unique elements of the array of prisms; and   c) determining if a conformational change was induced in the at least one target molecule through contact with the at least one test compound by detecting a change in a property of the nonlinear optical signal;   wherein determinations of conformational change are performed at an average rate of at least 10 test entities tested per hour.   
     
     
         2 . The method of  claim 1 , wherein the determinations of conformational change are performed at an average rate of at least 100 test compounds tested per hour. 
     
     
         3 . The method of  claim 1 , wherein the at least one target molecule is tethered to each of two or more discrete regions. 
     
     
         4 . The method of  claim 1 , wherein at least two different target molecules are tethered to at least two different discrete regions. 
     
     
         5 . The method of  claim 1 , wherein the target molecules are selected from the group consisting of cells, proteins, peptides, receptors, enzymes, antibodies, DNA, RNA, oligonucleotides, small molecules, and carbohydrates, or any combination thereof. 
     
     
         6 . The method of  claim 1 , wherein the target molecules are drug targets or portions thereof. 
     
     
         7 . The method of  claim 1 , wherein each of at least four different target molecules are tethered to at least four different discrete regions. 
     
     
         8 . The method of  claim 1 , wherein each discrete region comprises an area of up to about 100 mm 2  on the upper surface of the planar substrate. 
     
     
         9 . The method of  claim 7 , wherein each discrete region comprises a supported lipid bilayer. 
     
     
         10 . The method of  claim 9 , wherein the target molecules are tethered to or embedded within the supported lipid bilayer. 
     
     
         11 . The method of  claim 1 , wherein the at least one test compound is selected from the group consisting of cells, proteins, peptides, receptors, enzymes, antibodies, DNA, RNA, oligonucleotides, small molecules, and carbohydrates, or any combination thereof. 
     
     
         12 . The method of  claim 1 , wherein the test compound is a drug candidate or portion thereof. 
     
     
         13 . The method of  claim 1 , wherein the contacting step occurs in solution and comprises utilizing a pre-programmed fluid dispensing unit to dispense the at least one test compound. 
     
     
         14 . The method of  claim 1 , wherein the contacting step comprises contacting each of at least four different target molecules with a different test compound. 
     
     
         15 . The method of  claim 1 , wherein the contacting step comprises serially contacting the at least one target molecule with at least 100 different test compounds. 
     
     
         16 . The method of  claim 1 , wherein the contacting step comprises serially contacting the at least one target molecule with at least 10,000 different test compounds. 
     
     
         17 . The method of  claim 1 , wherein the one or more excitation light beams are provided by one or more lasers. 
     
     
         18 .- 19 . (canceled) 
     
     
         20 . The method of  claim 1 , wherein the non-linear optical signal is selected from the group consisting of second harmonic light, sum frequency light, and difference frequency light. 
     
     
         21 . The method of  claim 1 , further comprising moving said planar substrate relative to the position of one or more external sources of the one or more excitation light beams. 
     
     
         22 . The method of  claim 21 , wherein each discrete region of two or more discrete regions is optically coupled with an entrance prism and a different exit prism that are both integrated with the lower surface of the substrate. 
     
     
         23 .- 24 . (canceled) 
     
     
         25 . The method of  claim 1 , further comprising repeating the illuminating and determining steps a plurality of times after said contacting step, thereby determining conformational changes in the at least one target molecule as a function of time. 
     
     
         26 .- 42 . (canceled) 
     
     
         43 . The method of  claim 1 , wherein the change in a property of the nonlinear optical signal comprises a change in intensity, wavelength, or polarization.