Capture and detection format versatility for dipstick assays

Abstract

Dipsticks for testing for the presence of a target nucleic acid in a sample solution are described the dipsticks comprise a universal capture probe immobilised at a capture zone of the dipstick. The universal capture probe is capable of hybridising to a hook capture probe which is hybridised to the target nucleic acid in the sample solution. A contact end of the dipstick is contacted with the sample solution to cause hook capture probe hybridised to the target nucleic acid to move by capillary action to the capture zone where the target nucleic acid can be detected. Use of the universal and hook capture probes allows dipsticks to be prepared which can be used to capture any target nucleic acid, thereby simplifying preparation of the dipsticks. Specificity of target nucleic acid capture is then achieved by use of an appropriate hook capture probe. Methods and kits are also described.

1 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution and a universal capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end;   b) providing a hook capture probe that is capable of hybridizing to a first region of the target nucleic acid and to the universal capture probe;   c) incubating the sample solution with the hook capture probe under conditions for hybridization of the hook capture probe to the first region of the target nucleic acid in the sample solution;   d) contacting the sample solution with the contact end of the chromatographic strip so that a complex formed between the hook capture probe, the detection probe and the target nucleic acid can travel by capillary action to the capture zone and bind to the capture zone by interaction between the hook capture probe of the complex and the universal capture probe that is immobilized at the capture zone; and   e) detecting for target nucleic acid at the capture zone.   
     
     
         2 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution, a universal capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end and a hook capture probe that is releasably immobilized to the chromatographic strip between the contact end and the capture zone, wherein the hook capture probe is capable of hybridizing to a first region of the target nucleic acid and to the universal capture probe;   b) contacting the sample solution with the contact end of the chromatographic strip to cause sample solution to move by capillary action to the capture zone, thereby releasing the hook capture probe and allowing the released hook capture probe to hybridize to the target nucleic acid in the sample solution as it moves to the capture zone so that a complex comprising the target nucleic acid and the hook capture probe can be captured at the capture zone by hybridization of the universal capture probe to the hook capture probe of the complex; and   c) detecting for the target nucleic acid at the capture zone.   
     
     
         3 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution and a universal capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end;   b) providing a hook capture probe that is capable of hybridizing to a first region of the target nucleic acid and to the universal capture probe;   c) providing a detection probe capable of attaching to the target nucleic acid thereby allowing direct or indirect detection of the target nucleic acid utilizing the detection probe;   d) incubating the sample solution with the detection probe under conditions for attachment of the detection probe to the target nucleic acid in the sample solution and incubating the sample solution with the hook capture probe under conditions for hybridization of the hook capture probe to the first region of target nucleic acid in the sample solution;   e) contacting the sample solution with the contact end of the chromatographic strip so that a complex formed between the hook capture probe, the detection probe and target nucleic acid can travel by capillary action to the capture zone and bind to the capture zone by interaction between the hook capture probe of the complex and the universal capture probe that is immobilized at the capture zone; and   f) checking for the presence of the detection probe at the capture zone.   
     
     
         4 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution, a universal capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end, and a hook capture probe that is hybridized to the universal capture probe, wherein the hook probe is capable of hybridizing to a first region of the target nucleic acid;   b) contacting the sample solution with the contact end of the chromatographic strip so that the target nucleic acid in the sample solution can travel by capillary action to the capture zone and bind to the capture zone by interaction between the hook capture probe that is immobilized at the capture zone and the first region of the target nucleic acid; and   c) detecting for the target nucleic acid at the capture zone.   
     
     
         5 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution, a universal capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end, and a hook capture probe that is hybridized to the universal capture probe, wherein the hook probe is capable of hybridizing to a first region of the target nucleic acid;   b) providing a detection probe capable of attaching to the target nucleic acid thereby allowing direct or indirect detection of the target nucleic acid utilizing the detection probe;   c) incubating the sample solution with the detection probe under conditions for attachment of the detection probe to the target nucleic acid in the sample solution;   d) contacting the sample solution with the contact end of the chromatographic strip so that target nucleic acid attached to the detection probe can travel by capillary action to the capture zone and bind to the capture zone by interaction between the hook capture probe that is immobilized at the capture zone and the first region of the target nucleic acid; and   e) checking for the presence of the detection probe at the capture zone.   
     
     
         6 . The method according to  claim 5 , wherein the detection probe is capable of attaching to the target nucleic acid by hybridizing to a second region of the target nucleic acid. 
     
     
         7 . The method according to  claim 6 , wherein the detection probe comprises a hook detection probe capable of hybridizing to the second region of the target nucleic acid and a universal detection probe capable of hybridizing to the hook detection probe when the hook detection probe is hybridized to the target nucleic acid. 
     
     
         8 . The method according to  claim 7 , wherein the detection probe or the universal detection probe comprises a label thereby allowing direct detection of the target nucleic acid utilizing the detection probe or the universal and the hook detection probes. 
     
     
         9 . The method of  claim 7  further comprising contacting the detection probe or the universal detection probe with a detection moiety capable of binding by non base-pairing interaction to the detection probe when the detection probe is hybridized to the target nucleic acid, thereby allowing indirect detection of target nucleic acid utilizing the detection probe or the universal and the hook detection probes. 
     
     
         10 . The method according to  claim 9 , wherein the detection probe or the universal detection probe comprises a detection ligand which can bind the detection moiety. 
     
     
         11 . The method according to  claim 9 , wherein the detection moiety comprises an antibody or an antibody fragment. 
     
     
         12 . The method according to  claim 10 , wherein the detection ligand comprises biotin and the detection moiety comprises an anti-biotin antibody, or wherein the detection ligand comprises fluorescein and the detection moiety comprises an anti-fluorescein antibody, or wherein the detection ligand comprises 2,4-dinitrophenol (DNP) and the detection moiety comprises an anti-DNP antibody. 
     
     
         13 . The method according to  claim 9 , wherein the detection moiety is releasably immobilized at a conjugate zone located between the contact end and the capture zone of the chromatographic strip so that the detection moiety is released and can bind to the detection probe as the complex travels by capillary action to the capture zone. 
     
     
         14 . The method according to  claim 9 , wherein the detection moiety comprises a label allowing detection of the detection moiety. 
     
     
         15 . The method according to  claim 8 , wherein the label is a non radioactive label. 
     
     
         16 . The method according to  claim 15 , wherein the label is a color label. 
     
     
         17 . The method according to  claim 16 , wherein the color label comprises a textile dye, a metal sol such as colloidal gold, or colored particles such as colored latex particles. 
     
     
         18 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution and a capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end, wherein the capture probe is capable of hybridizing to a first region of the target nucleic acid;   b) providing a hook detection probe that is capable of hybridizing to a second region of the target nucleic acid;   c) providing a universal detection probe that is capable of hybridizing to the hook detection probe when the hook detection probe is hybridized to the second region of the target nucleic acid, thereby allowing direct or indirect detection of target nucleic acid utilizing the universal and the hook detection probes;   d) incubating the sample solution with the hook detection probe and the universal detection probe under conditions for hybridization of the hook detection probe to the second region of the target nucleic acid and for hybridization of the universal detection probe to the hook detection probe;   e) contacting the sample solution with the contact end of the chromatographic strip so that a complex formed between the universal detection probe, the hook detection probe and the target nucleic acid can travel by capillary action to the capture zone and bind to the capture zone by interaction between the target nucleic acid of the complex and the capture probe that is immobilized at the capture zone; and   f) checking for the presence of the universal detection probe at the capture zone.   
     
     
         19 . A method for testing for the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution, a capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end, wherein the capture probe is capable of hybridizing to a first region of the target nucleic acid and a universal detection probe that is releasably immobilized to the chromatographic strip between the contact end and the capture zone;   b) incubating the sample solution with a hook detection probe that is capable of hybridizing to a second region of the target nucleic acid, under conditions for hybridization of the hook detection probe to the second region;   c) contacting the sample solution with the contact end of the chromatographic strip to cause sample solution to travel by capillary action to the capture zone, thereby releasing the universal detection probe so that the released universal detection probe can hybridize to the hook detection probe in the sample solution as it travels to the capture zone and a complex comprising the universal detection probe, the hook detection probe and the target nucleic acid can bind to the capture zone by interaction between the target nucleic acid of the complex and the capture probe that is immobilized at the capture zone; and   f) checking the presence of the universal detection probe at the capture zone.   
     
     
         20 . A method for testing the presence of a target nucleic acid in a sample solution comprising:
 a) providing a chromatographic strip having a contact end for contacting the sample solution and a capture probe that is immobilized at a capture zone of the chromatographic strip remote from the contact end, wherein the capture probe is capable of hybridizing to a first region of the target nucleic acid;   b) providing a hook detection probe that is capable of hybridizing to a second region of the target nucleic acid;   c) providing a universal detection probe that is capable of hybridizing to the hook detection probe when the hook detection probe is hybridized to the second region of the target nucleic acid, thereby allowing direct or indirect detection of the target nucleic acid utilizing the universal and the hook detection probes;   d) contacting the sample solution with the contact end of the chromatographic strip to cause sample solution to travel by capillary action to the capture zone and be captured at the capture zone by the capture probe;   e) contacting the capture zone with the hook detection probe and the universal detection probe; and   f) checking the presence of the universal detection probe at the capture zone.   
     
     
         21 . The method according to  claim 18 , wherein the universal detection probe comprises a label. 
     
     
         22 . The method of  claim 20  further comprising contacting the universal detection probe with a detection moiety capable of binding by non base pairing interaction to the universal detection probe when the universal detection probe is hybridized to the hook detection probe. 
     
     
         23 . The method according to  claim 22 , wherein the universal detection probe comprises a detection ligand which can be bound by the detection moiety. 
     
     
         24 . The method according to  claim 22 , wherein the detection moiety comprises an antibody or an antibody fragment. 
     
     
         25 . The method according to  claim 24 , wherein the detection ligand comprises a biotin and the detection moiety comprises an anti-biotin antibody, or wherein the detection ligand comprises a fluorescein and the detection moiety comprises an anti-fluorescein antibody, or wherein the detection ligand comprises a 2,4-dinitrophenol (DNP) and the detection moiety comprises an anti-DNP antibody. 
     
     
         26 . The method according to  claim 22 , wherein the detection moiety is releasably immobilized at a conjugate zone located between the contact end and the capture zone of the chromatographic strip so that the detection moiety is released and can bind to the universal detection probe as the complex travels by capillary action to the capture zone. 
     
     
         27 . A method for testing the presence of a target nucleic acid in a sample solution comprising: 
     
     
         28 - 63 . (canceled)