Methods for diagnosing and treating cerebrovascular events based on NR2 peptides
Methods and kits for diagnosing and treating cerebrovascular events, and for defining the time and anatomical location of an event, are provided based on the detection and quantification of bound or total and unbound NR2 peptides in biological fluids. The methods are optionally performed in conjunction with neurological scoring and neuroimaging, and are directed to risk assessment, prognosis, diagnosis and treatment of TIA and stroke on an emergency basis in the emergency room.
1 ) A method for determining the pathological origin of a neurological deficit comprising:
a) providing a patient suffering from a neurological deficit;
b) detecting and quantifying bound or total NR2 peptide in a biological fluid from said patient to arrive at a quantity of bound or total NR2 peptide; and
c) comparing said quantity of bound or total NR2 peptide with a population norm for bound or total NR2 peptide in said biological fluid in apparently healthy human subjects.
2 ) The method of claim 1 wherein said population norm for said total NR2 peptide in blood is about 2.0 ng/ml. or less.
3 ) The method of claim 1 further comprising:
a) detecting and quantifying unbound NR2 peptide in a biological fluid from said patient to arrive at a quantity of unbound NR2 peptide; and
b) comparing said quantity of unbound NR2 peptide with a population norm for unbound NR2 peptide in said biological fluid in apparently healthy human subjects.
4 ) The method of claim 3 wherein said population norm for said unbound NR2 peptide in blood is less than about 1.0 ng/ml.
5 ) The method of claim 3 , further comprising:
a) calculating a total:unbound NR2 peptide ratio by dividing said quantity of said total NR2 peptide by said quantity of said unbound NR2 peptide;
b) correlating a total:unbound NR2 peptide ratio less than 2:1 and a quantity of unbound NR2 peptide greater than 1.0 ng./ml. with a primary neurovascular event; and
c) correlating a total:unbound NR2 peptide ratio greater than 2:1 and a quantity of total NR2 peptide greater than 2.0 ng./ml. with a recurrent neurovascular event.
6 ) The method of claim 3 , further comprising correlating a quantity of total NR2 peptide of less than 2.0 ng/ml and a quantity of unbound NR2 peptide of less than 1.0 ng/ml with the absence of an ischemic event in the cerebrovasculature.
7 ) The method of claim 3 further comprising performing steps (b) and (c) one or more additional times, and correlating changes in said quantity of unbound NR2 peptide to a neurological event, and a constant quantity of unbound NR2 peptide at levels below said population norm to a lack of a neurological event.
8 ) The method of claim 1 , further comprising neuroimaging the brain of said patient by computed tomography or magnetic resonance/diffusion weighted imaging.
9 ) The method of claim 1 wherein said biological fluid is blood, urine, blood plasma, blood serum, cerebrospinal fluid, saliva, perspiration or brain tissue, or a derivative thereof.
10 ) A method for predicting the likelihood of a cerebrovascular event comprising:
a) providing a patient at risk for suffering a cerebrovascular event;
b) detecting and quantifying bound or total NR2 peptide in a biological fluid from said patient to arrive at a quantity of bound or total NR2 peptide; and
c) comparing said quantity of bound or total NR2 peptide with a population norm for bound or total NR2 peptide in said biological fluid in apparently healthy human subjects.
11 ) The method of claim 10 wherein said population norm for said total NR2 peptide in blood is about 2.0 ng/ml. or less.
12 ) The method of claim 10 further comprising:
a) detecting and quantifying unbound NR2 peptide in a biological fluid from said patient to arrive at a quantity of unbound NR2 peptide; and
b) comparing said quantity of unbound NR2 peptide with a population norm for unbound NR2 peptide in said biological fluid in apparently healthy human subjects.
13 ) The method of claim 12 wherein said population norm for said unbound NR2 peptide in blood is less than about 1.0 ng/ml.
14 ) The method of claim 10 wherein said patient is scheduled for surgery.
15 ) A test kit for detecting and quantifying bound or total NR2 peptide in a biological sample comprising:
a) an antibody reagent comprising antibodies that are capable of specifically binding NR2 peptide in a test sample;
b) a protein detergent; and
c) an indicator reagent.
16 ) The test kit of claim 15 further comprising synthetic NR2 peptide as a control or calibrator.
17 ) A method for detecting and quantifying bound or total NR2 peptide in a biological sample comprising:
a) contacting a biological sample with a protein detergent to yield a denatured biological sample;
b) contacting said denatured biological sample with an antibody reagent comprising antibodies that are capable of specifically binding NR2 peptide in a test sample, for a time sufficient to allow formation of bound complexes between said NR2 peptide and said antibody; and
c) detecting and quantitating said bound complexes.
18 ) The method of claim 17 , wherein the biological sample is plasma diluted to a ratio of about 1:50.
19 ) The method of claim 17 , wherein said detergent is 0.1-0.5% sodium dodecyl sulphate or KCl.
20 ) The method of claim 17 wherein:
a) said antibody reagent comprises antibodies bound to an agglutinating carrier; and
b) said complexes are detected and measured based on the degree of agglutination.
21 ) The method of claim 17 , wherein the sufficient time period is 5 minutes or less.
22 ) The method of claim 17 performed by direct ELISA, RIA, immunodot, immunoblot, latex agglutination, lateral flow, fluorescence polarization assay, or microarray.
23 ) The method of claim 17 , wherein said detecting and quantitating comprises:
a) generating a signal from a signal generating compound by:
i) contacting said complex with a second antibody reagent comprising IgG linked to said signal generating reagent to form a second complex, or
ii) contacting said complex with a second antibody reagent comprising IgG to form a second complex, and subsequently linking said signal generating reagent to said IgG; and
b) measuring the signal generated; wherein the amount of signal measured correlates to the amount of said NR2 present in said sample.
24 ) The method of claim 17 , wherein said IgG comprises chicken anti-human or anti-human IgG and said indicator reagent comprises horseradish peroxidase.
25 ) A method of diagnosing and treating cerebral ischemia comprising:
a) providing a patient suffering from a neurological deficit;
b) detecting and quantifying NR2 peptide in a biological fluid from said patient to obtain a first quantity of NR2 peptide;
c) comparing said first quantity of NR2 peptide with a population norm for NR2 peptide in said biological fluid in apparently healthy human subjects;
d) administering tPA to said patient if said first quantity of said NR2 peptide is above the population norm for NR2 peptide in said biological fluid in apparently healthy human subjects;
e) not administering tPA to said patient if said quantity of said NR2 peptide is below the population norm for NR2 peptide in said biological fluid in apparently healthy human subjects.
26 ) The method of claim 25 wherein bound and unbound NR2 peptide are separately detected and quantified to arrive at quantities of bound or total and unbound NR2 peptides, and said quantities of said bound or total and unbound NR2 peptide are compared with population norms of bound or total and unbound NR2 peptides in said biological fluid in apparently healthy human subjects.
27 ) The method of claim 25 wherein said population norm for said bound NR2 peptide in blood is less than about 2.0 ng/ml.
28 ) The method of claim 25 wherein said population norm for said unbound NR2 peptide in blood is less than about 1.0 ng/ml.
29 ) The method of claim 25 further comprising:
a) evaluating said patient for acute facial paresis, arm drift, or abnormal speech;
b) administering tPA to said patient if said acute facial paresis, arm drift, or abnormal speech is observed and said quantity of said NR2 peptide is above the population norm for NR2 peptide in said biological fluid in apparently healthy human subjects; and
c) not administering tPA to said patient if said acute facial paresis, arm drift, or abnormal speech is not observed and said quantity of said NR2 peptide is below the population norm for NR2 peptide in said biological fluid in apparently healthy human subjects.
30 ) The method of claim 25 , further comprising:
a) detecting and quantifying NR2 peptide in a biological fluid from said patient a second time to obtain a second quantity of NR2 peptide;
b) comparing said second quantity of NR2 peptide to said first quantity; and
c) administering tPA a second time to said patient if the second quantity is greater than the first quantity; and
d) not administering tPA a second time to said patient if said second quantity is not greater than the first quantity.