Method for labeling and detecting materials employing luminescent arysulfonate cyanine dyes

A method for labeling a first component and using the labeled first component to bind with a second component to form a labeled second component, said method comprising:

• (a) adding to an aqueous liquid containing a first component selected from the group consisting of antibodies, antigens and nucleic acids, a luminescent dye selected from the group consisting of cyanine, merocyanine and styryl dyes having the structures:
• ##STR11## wherein:
• the dotted lines each represent carbon atoms necessary for the formulation of one ring to three fused rings having 5 to 6 carbon atoms in each ring;
• X and Y are selected from the group consisting of O, S, and CH.sub.3 --C--CH.sub.3 ;
• m is an integer selected from the group consisting of 1, 2, 3 and 4;
• at least one of said R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5, R.sub.6 and R.sub.7 groups contains at least one reactive group to make said dye covalently reactive with said first component and,
• at least one of said R.sub.8 and R.sub.9 groups comprises a sulfonic acid or sulfonate group attached to an aromatic nucleus of said dye;
• (b) reacting said dye with said first component so that said dye covalently binds to and luminescently labels said first component to provide a labeled first component;
• (c) treating a sample with said labeled first component for binding said labeled first component to a second component, if any, present in said sample, said second component having at least a portion comprised of an antigen, antibody or complementary nucleic acid capable of binding to said respective antibody, antigen or nucleic acid of said labeled first component to form a labeled second component;
• and
• (d) detecting said labeled second component by an optical method.

A method for labeling a component of an aqueous liquid comprising:

• (a) adding to a liquid which contains a component selected from the group consisting of proteins, sugars, cells, nucleic acids and combinations thereof, a luminescent dye covalently reactive with an amine, aldehyde, sulfhydryl or hydroxy group on said component and at least one sulfonic acid or sulfonate group attached to an aromatic nucleus of said dye;
• wherein said dye is selected from the group consisting of:
• ##STR12## wherein:
• the dotted lines each represent carbon atoms necessary for the formulation of one ring to three fused rings having 5 to 6 carbon atoms in each ring;
• X and Y are selected from the group consisting of O, S, ##STR13## m is an integer selected from the group consisting of 1, 2, 3 and 4;
• at least one of said R.sub.1, R.sub.2, R.sub.3, R.sub.4, R.sub.5, R.sub.6 and R.sub.7 groups contains at least one moiety selected from the group consisting of isothiocyanate, isocyanate, monochlorotriazine, dichlorotriazine, mono- or di-halogen substituted pyridine, mono or di-halogen substituted diazine, maleimide, aziridine, azidonitrophenyl, azide, 3-(2-pyridyl dithio)-proprionamide, glyoxal and aldehyde;
• and
• at least one of said R.sub.8 and R.sub.9 groups comprises at least one arylsulfonyl or arylsulfonate moiety;
• and
• (b) reacting said dye with said component so that said dye covalently binds to and labels said component.

A method comprising:

• (a) in a liquid which contains a first component, at least a portion of said first component being selected from the group consisting of proteins, sugars, nucleic acids, cells and combinations thereof, and having an amino, hydroxy or sulfhydryl nucleophile, labeling said first component with a water soluble luminescent dye comprising a cyanine having the structure ##STR16## wherein:
• the dotted lines each represent carbon atoms necessary for the formation of said dye;
• X and Y are each selected from the group consisting of O, S and CH.sub.3 --C--CH.sub.3 ;
• m is an integer selected from the group consisting of 1, 2 and 3;
• at least one of said R.sub.1, R.sub.2, R.sub.3, R.sub.4, and R.sub.7 group reacts with amino, hydroxy or sulfhydryl nucleophiles of said first component and is a reactive moiety selected from a group containing ##STR17## wherein Q is Br or Cl, and n=0, 1, 2, 3, 4, 5, 6, 7 or 8 for R.sub.3, R.sub.4, and R.sub.7, and n=1, 2, 3, 4, 5, 6, 7 or 8 for R.sub.1 and R.sub.2 ;
• and when any one of said R.sub.3, R.sub.4, and R.sub.7 groups is not selected from one said reactive moiety, said remaining R.sub.3, R.sub.4, and R.sub.7 are hydrogen or E-F, a polar group wherein E is a spacer group having the structure --(CH.sub.2).sub.n -- and n of said spacer group=0, 1, 2, 3, or 4, and F is hydroxy, sulfonate, sulfate, carboxylate, or a lower alkyl substituted amino;
• and when any one of said R.sub.1 and R.sub.2 groups is not selected from one said reactive moiety, said remaining R.sub.1 and R.sub.2 are a lower alkyl or said E-F and n of said spacer group=1, 2, 3, or 4;
• at least one said R.sub.8 and R.sub.9 groups is selected from the group consisting of an arylsulfonic acid and an arylsulfonate moiety for conferring improved luminescence to said dye when in use as compared to a dye which does not have said at least one arylsulfonic acid or arylsulfonate moiety;
• and when one of said R.sub. or R.sub.9 group is not an arylsulfonic acid or an arylsulfonate, said remaining R.sub.8 or R.sub.9 group is a hydrogen;
• (b) contacting said labeled first component with a second component having at least a portion thereof capable of binding to said first component to form a labeled second component;
• and,
• (c) detecting the presence of said labeled first and second components by optical methods.