Method for detecting particulate substance by immunochromatography, and kit for the same
An object of the present invention is to provide a method for detecting particulate substances with a high degree of sensitivity, while suppressing the generation of non-specific signals. According to the present invention, the generation of non-specific signals in the background can be suppressed by blocking a membrane of a test strip which is used for immunochromatography with a blocking composition including a polymer-based blocking agent having, in its side chain, a substituent represented by the following formula: (wherein R 1 , R 2 , and R 3 are each independently a hydrogen atom, a C 1-6 alkyl group, or a C 1-6 hydroxyalkyl group; and n is an integer of 1 to 4), and having a weight average molecular weight (Mw) of 1×10 3 to 1×10 7 .
1 . A method for detecting a particulate substance contained in various body fluids, including breast milk, saliva, and tears, as well as blood and urine; preservative solutions for tissue sections; or cell culture supernatants by immunochromatography, the particulate substance comprising, on its surface, a plurality of substances to be bound including a first substance to be bound and a second substance to be bound that may be the same or different from each other, the method comprising steps of:
providing a test strip comprising a membrane on which a first specific binding substance for the first substance to be bound is immobilized, wherein the membrane is blocked with a blocking composition comprising a polymer-based blocking agent having, in its side chain, a substituent represented by the following formula:
(wherein R 1 , R 2 , and R 2 are each independently a hydrogen atom, a C 1-6 alkyl group, or a C 1-6 hydroxyalkyl group; and n is an integer of 1 to 4), and having a weight average molecular weight (Mw) of 1×10 3 to 1×10 7 ;
providing a second specific binding substance for the second substance to be bound, the second specific binding substance binding to a labeling substance;
contacting a sample containing the particulate substance with the first specific binding substance to capture the particulate substance on the membrane;
contacting the particulate substance with the second specific binding substance to label the particulate substance;
detecting the labeled particulate substance captured on the membrane.
2 . The method according to claim 1 , wherein the particulate substance comprises extracellular vesicles.
3 . The method according to claim 2 , wherein the extracellular vesicles are exosomes.
4 . The method according to claim 1 , wherein the detecting step is performed by measuring chemiluminescence or fluorescence.
5 . The method according to claim 1 , wherein the labeling step is performed in the presence of the polymer-based blocking agent.
6 . The method according to claim 1 , wherein the polymer-based blocking agent is LIPIDURE®.
7 . The method according to claim 1 , wherein the blocking composition further comprises an additional blocking agent.
8 . A method for producing a kit for detecting a particulate substance contained in various body fluids, including breast milk, saliva, and tears, as well as blood and urine; preservative solutions for tissue sections; or cell culture supernatants by immunochromatography, the particulate substance comprising, on its surface, a plurality of substances to be bound including a first substance to be bound and a second substance to be bound that may be the same or different from each other, the method comprising steps of:
providing a test strip comprising a membrane on which a first specific binding substance for the first substance to be bound is immobilized;
providing, in order to block the membrane, a blocking composition comprising a polymer-based blocking agent having, in its side chain, a substituent represented by the following formula:
(wherein R 1 , R 2 , and R 2 are each independently a hydrogen atom, a C 1-6 alkyl group, or a C 1-6 hydroxyalkyl group; and n is an integer of 1 to 4), and having a weight average molecular weight (Mw) of 1×10 3 to 1×10 7 ;
blocking the membrane with the blocking composition.
9 . The production method according to claim 8 , further comprising steps of:
providing a second specific binding substance for the second substance to be bound, the second specific binding substance binding to a labeling substance; and
blocking the second specific binding substance with the blocking composition.
10 . A kit for detecting a particulate substance contained in various body fluids, including breast milk, saliva, and tears, as well as blood and urine; preservative solutions for tissue sections; or cell culture supernatants by immunochromatography, the particulate substance comprising, on its surface, a plurality of substances to be bound including a first substance to be bound and a second substance to be bound that may be the same or different from each other, the kit comprising:
a test strip comprising a membrane on which a first specific binding substance for the first substance to be bound is immobilized, wherein the membrane is blocked with a blocking composition comprising a polymer-based blocking agent having, in its side chain, a substituent represented by the following formula:
(wherein R 1 , R 2 , and R 2 are each independently a hydrogen atom, a C 1-6 alkyl group, or a C 1-6 hydroxyalkyl group; and n is an integer of 1 to 4), and having a weight average molecular weight (Mw) of 1×10 3 to 1×10 7 ;
a second specific binding substance for the second substance to be bound, the second specific binding substance binding to a labeling substance.