Fluorescent Dyes, Fluorescent Dye Kits, and Methods of Preparing Labeled Molecules

Abstract

The present invention provides methods, compositions, and kits useful in preparing labeled molecules, which are useful in the detection of binding partners.

1 - 104 . (canceled) 
     
     
         105 . A method of preparing a labeled protein, comprising:
 (a) providing an amine-reactive detectable dye or an amine-reactive detectable label;   (b) combining said amine-reactive detectable dye or said amine-reactive detectable label, in a buffer, with a sample solution comprising a target protein, said target protein comprising an amine; and   (c) allowing said amine reactive detectable dye or said amine-reactive detectable label to react with said amine in said buffer for about 60 minutes or less;   thereby producing said labeled protein, wherein said labeled protein is labeled with said amine-reactive detectable dye or said amine-reactive detectable label.   
     
     
         106 . The method of  claim 105 , wherein the reaction of step (c) is completed in about 45 minutes or less. 
     
     
         107 . The method of  claim 105 , wherein the reaction of step (c) is completed in about 30 minutes or less. 
     
     
         108 . The method of  claim 105 , wherein the reaction of step (c) is completed in about 20 minutes or less. 
     
     
         109 . The method of  claim 105 , wherein said amine-reactive dye or said amine-reactive detectable label comprises one or more activated esters. 
     
     
         110 . The method of  claim 109 , wherein said one or more activated esters is an N-hydroxy succinimidyl ester, N-hydroxy sulfosuccinimidyl ester, p-sulfo-tetrafluorophenol ester, or a combination thereof 
     
     
         111 . The method of  claim 110 , wherein said one or more activated esters is an N-hydroxy succinimidyl ester. 
     
     
         112 . The method of  claim 105 , wherein said amine-reactive detectable dye or said amine-reactive detectable label comprises one or more sulfonate groups. 
     
     
         113 . The method of  claim 105 , wherein said sample solution is not pretreated before steps (a) or (b), wherein said pretreatment comprises purification of said target protein away from one or more other components of said sample solution. 
     
     
         114 . The method of  claim 113 , wherein said purification step comprises protein A-based chromatography, size-exclusion chromatography, or ultra membrane filtration, or a combination thereof. 
     
     
         115 . The method of  claim 105 , wherein said target protein is an antibody or a fragment thereof. 
     
     
         116 . The method of  claim 105 , wherein said amine-reactive detectable dye or said amine-reactive detectable label is directly observable. 
     
     
         117 . The method of  claim 105 , wherein said amine-reactive detectable dye or said amine-reactive detectable label is a fluorescent dye. 
     
     
         118 . The method of  claim 105 , wherein said amine-reactive detectable dye or said amine-reactive detectable label comprises a biotin. 
     
     
         119 . The method of  claim 105 , wherein said amine-reactive detectable dye is a CF dye, an Alexa Fluor dye, a DyLight Dye, a Cy dye, an IRDye, a HiLyte dye, a sulfonated or pegylated coumarin dye, a sulfonated or pegylated xanthene dye, a sulfonated or pegylated cyanine dye, or a sulfonated or pegylated pyrene dye. 
     
     
         120 . The method of  claim 105 , wherein said sample solution further comprises one or more stabilizers or preservatives. 
     
     
         121 . The method of  claim 120 , wherein at least one of said one or more stabilizers or preservatives is bovine serum albumin, gelatin, or glycerol. 
     
     
         122 . The method of  claim 121 , wherein at least one of said one or more stabilizers or preservatives is bovine serum albumin. 
     
     
         123 . The method of  claim 121 , wherein at least one of said one or more stabilizers or preservatives is glycerol. 
     
     
         124 . The method of  claim 120 , wherein at least one of said one or more stabilizers or preservatives is sodium azide.