Colorimetric Labeling and Detection Methods and Compositions
AssigneeLife Technologies Corporation
InventorsScott CLARKE, Kyle GEE, Carmen FINNESSY
Life of this patent
Estimating the term…
Abstract
The present disclosure provides methods, compositions and kits for measuring cellular nascent nucleic acid synthesis by colorimetric labeling of nucleic acid. The nucleic acid synthesis can be measured as cell proliferation, DNA or RNA synthesis, gene expression or apoptosis. Additionally, these methods may be used to screen compounds for their effect on cellular proliferation by treating cells or an organism with the test compound simultaneous to or before treatment with a competitive nucleoside analog.
Claims24
1-58. (canceled)
A method of measuring cellular proliferation in an organism, the method comprising:
- a) administering to an organism an effective amount of an alkynyl-modified nucleoside analogue, such that the alkynyl-modified nucleoside analogue is incorporated into DNA of cells of the organism;
- b) contacting at least one cell of the organism with an azide-modified biotin under conditions such that the alkynyl moiety of the alkynyl-modified nucleoside analogue forms a covalent link with the azide moiety of the biotin;
- c) contacting the at least one cell of the organism with a (strept)avidin conjugated to a detectable label;
- and
- d) measuring the amount of detectable label, wherein the amount of label indicates the extent of cellular proliferation.
(canceled)
A method for identifying an agent that perturbs cellular proliferation in an organism, the method comprising:
- a) exposing an organism to a test agent;
- b) administering to the organism an effective amount of an alkynyl-modified nucleoside analogue, such that the alkynyl-modified nucleoside analogue is incorporated into DNA of cells of the organism;
- c) contacting at least one cell of the organism with an azide-modified biotin under conditions such that the alkynyl moiety of the alkynyl-modified nucleoside analogue forms a covalent link with the azide moiety of the biotin;
- d) contacting the at least one cell of the organism with a (strept)avidin conjugated to a detectable label;
- e) measuring the amount of detectable label, wherein the amount of label indicates the extent of cellular proliferation;
- and
- f) identifying the test agent as an agent that perturbs cellular proliferation if the amount of label measured in step (e) is less than or greater than the amount of label measured in a control application in which the organism is not exposed to the test agent.
62-63. (canceled)
A method of detecting apoptosis, the method comprising:
- a) contacting a cell with an effective amount of an alkynyl-modified nucleotide analogue and a terminal deoxynucleotidyl transferase (TdT), such that the alkynyl-modified nucleotide analogue is incorporated into DNA of the cell;
- b) contacting the cell with an azide-modified biotin under conditions such that the alkynyl moiety of the alkynyl-modified nucleotide analogue forms a covalent link with the azide moiety of the biotin;
- c) contacting the cell with a (strept)avidin conjugated to a detectable label;
- and
- d) measuring the amount of detectable label, wherein the amount of label indicates the amount of apoptosis.
(canceled)
Dependencies are inferred from the claim text.