Characterization of reaction variables

Abstract

A microscale method for the characterization of one or more reaction variables that influence the formation or dissociation of an affinity complex comprising a ligand and a binder, which have mutual affinity for each other. The method is characterized in comprising the steps of: (i) providing a microfluidic device comprising a microchannel structures that are under a common flow control, each microchannel structure comprising a reaction microactivity; (ii) performing essentially in parallel an experiment in each of two or more of the plurality of microchannel structures, the experiment in these two or more microchannel structures comprising either a) formation of an immobilized form of the complex and retaining under flow conditions said form within the reaction microactivity, or b) dissociating, preferably under flow condition, an immobilized form of the complex which has been included in the microfluidic device provided in step (i), at least one reaction variable varies or is uncharacterized for said two or more microchannel structures while the remaining reaction variables are kept essentially constant; (iii) measuring the presentation of the complex in said reaction microactivity in said two or more microchannel structures; and (iv) characterizing said one or more reaction variables based on the values for presentation obtained in step (iii).

1 . A microscale method for the characterization of at least one reaction variable that influences the dissociation of an affinity complex comprising a ligand and a binder, which have affinity for each other, said method comprising the steps of:
 (i) providing a microfluidic device comprising a plurality of microchannel structures that are under common flow control, each microchannel structure comprising a reaction microcavity;   (ii) performing essentially in parallel an experiment in each of two or more of the plurality of microchannel structures, the experiment in these two or more microchannel structures comprising
 dissociating under flow conditions the immobilized form of the complex which has been introduced prior to step (ii), 
 said at least one reaction variable being different or uncharacterized for said two or more microchannel structures; 
   (iii) measuring the presentation of the complex in said reaction microcavity in each of said two or more microchannel structures; and   (iv) characterizing said at least one reaction variable based on the values for presentation obtained din step (iii).   
     
     
         2 . The method of  claim 1 , wherein each of the microcavities of said two or more microchannel structures comprises a solid phase to which an affinity reactant which is capable of being incorporated into the affinity complex retained in step (ii) is attached. 
     
     
         3 . The method of  claim 1 , wherein
 (a) the microfluidic device comprises a substrate having an axis of symmetry,   (b) each microchannel structure is oriented relative the axis of symmetry with an inlet port at shorter radial distance than the reaction microcavity, and   (c) the substrate is spun around its axis of symmetry to drive liquid within the microchannel structures.   
     
     
         4 . The method of  claim 1 , wherein
 a) the microfluidic device comprises a substrate having an axis of symmetry,   b) each microchannel structure is oriented radially relative the axis of symmetry with the reaction microcavity at a larger radial distance than a substructure delivering liquid to the reaction microcavity, and   c) the substrate is spun around its axis of symmetry to drive liquid within the microchannel structures.   
     
     
         5 . The method of  claim 1 , wherein each of the microchannel structures comprises a flow restriction downstream the reaction microcavity, which creates a pressure drop that restricts the flow through the reaction microcavity. 
     
     
         6 . The method of  claim 1 , wherein step (iii) is performed by measuring
 (a) distribution of the complex in the reaction microcavity along the flow direction, or   (b) the total amount of the complex in the reaction microcavity.   
     
     
         7 . The method of  claim 1 , wherein step (iii) comprises measuring the total amount of the complex in the reaction microcavity. 
     
     
         8 . The method of  claim 1 , wherein each of said experiments comprises formation of an immobilized form of the complex within the reaction microcavity. 
     
     
         9 . The method of  claim 1 , wherein said experiment comprises dissociating under flow conditions an immobilized form of the complex which complex is included in the microfluidic device provided in step (i). 
     
     
         10 . The method of  claim 1 , wherein said at least one reaction variables that varies between the microchannel structure is selected amongst group 1 or group 2 reaction variables. 
     
     
         11 . The method of  claim 1 , wherein the formation of the complex that is retained in the reaction microcavity is part of a competitive/inhibition protocol or a non-competitive protocol. 
     
     
         12 . The method of  claim 11 , wherein said at least one reaction variable is related to an affinity reactant. 
     
     
         13 . The method of  claim 12 , wherein said at least one reaction variable is related to the amount of said affinity reactant. 
     
     
         14 . The method of  claim 12 , wherein said at least one reaction variable is related to affinity of an affinity reactant. 
     
     
         15 . The method of  claim 1 , wherein step (iii) comprises determining the distribution of the complex along the flow direction in the reaction microcavity in each of said two or more microchannel structures. 
     
     
         16 . The method of  claim 15 , wherein step (iv) comprises that the relative affinity for the ligand and binder as a function of the variation in said at least one reaction variable is characterized.