Conjugates of 1,4,7-triazacyclononanes, dinuclear metal complexes of such conjugates, and methods of use for both 1,4,7-triazacyclononanes and conjugates

A compound having the formula

• in which:
• one or more of R1 through R6 is -L-R7 in which L is a linking group and R7 is a member selected from the group consisting of an acrylamide group, a fluorescent dye, an affinity-type binding member, and a solid chromatographic support;
• and
• the remainder of R1 through R6 are independently selected from the group consisting of H and C1-C6 alkyl.

A compound having the formula

• in which:
• one of R1 through R6 is -L-R7 in which L is a linking group and R7 is a member selected from the group consisting of an acrylamide group, a fluorescent dye, an affinity-type binding member, and a solid chromatographic support;
• the remainder of R1 through R6 are independently selected from the group consisting of H and C1-C6 alkyl;
• and
• M is a divalent metal.

A method for the preparation of a separation medium for use in electrophoresis of phosphorylated species, said method comprising polymerizing a monomer mixture comprising acrylamide, a crosslinker, an initiator, and a compound having the formula

• in which:
• one of R1 through R6 is -L-R7 in which L is a linking group and R7 is an acrylamide group;
• the remainder of R1 through R6 are independently selected from the group consisting of H and C1-C6 alkyl;
• and
• M is a divalent metal.

A method of staining phosphorylated compounds immobilized on a solid support or in a gel with a fluorescent dye, said method comprising contacting said solid support or gel with a compound having the formula

• in which:
• one of R1 through R6 is -L-R7 in which L is a linking group and R7 is a fluorescent dye;
• the remainder of R1 through R6 are independently selected from the group consisting of H and C1-C6 alkyl;
• and
• M is a divalent metal.

A method for detecting a phosphorylated compound immobilized on a solid support or in a gel, said method comprising:

• (A) contacting said solid support or gel with a compound having the formula
• in which:
• one of R1 through R6 is -L-R7 in which L is a linking group and R7 is a first binding member;
• the remainder of R1 through R6 are independently selected from the group consisting of H and C1-C6 alkyl;
• and
• M is a divalent metal;
• and
• (B) subsequent to (A), contacting said solid support or gel with a conjugate of (i) a second binding member that binds to said first binding member upon contact through affinity type binding and (ii) a reporter group.

A method for extraction of phosphorylated species from a liquid solution, said method comprising passing said liquid solution through an extraction medium comprising the following compound conjugated to a solid support:

• in which:
• one of R1 through R6 is -L-* in which L is a linking group and * is a site of attachment to said solid support;
• the remainder of R1 through R6 are independently selected from the group consisting of H and C1-C6 alkyl;
• and
• M is a divalent metal.