Method of detecting human β-defensins
A method of detecting β-defensin in a bodily sample from a subject includes reducing the electrostatic interaction between β-defensin and negatively charged moieties in the bodily sample prior to detecting the β-defensin with an antibody or epitope binding fragment thereof.
1. A method of detecting β-defensin in a bodily sample of a subject comprising:
obtaining a bodily sample from the subject;
reducing the electrostatic interaction between β-defensin and negatively charged moieties in the bodily sample;
contacting the bodily sample with a capture antibody or an epitope binding fragment thereof, the capture antibody or an epitope binding fragment thereof being immunoreactive with and binding to the β-defensin; and
detecting bound β-defensin in the bodily sample.
2. The method of claim 1 , the step of detecting bound β-defensin captured comprising:
contacting the bound β-defensin with a detection antibody or epitope binding fragment thereof, the detection antibody or epitope binding fragment thereof being coupled to a detectable label; and
detecting the detectable label in the bodily sample.
3. The method of claim 1 , the step of reducing the electrostatic interaction between β-defensin and negatively charged moieties in the sample comprising adding positively charged moieties to the bodily sample at an amount effective to reduce the electrostatic interactions between β-defensin and negatively charged moieties in the bodily sample.
4. The method of claim 3 , the positively charged moieties comprising cations.
5. The method of claim 4 , the cations being selected from the group consisting of Mg 2+ , Ca 2+ , and combinations thereof.
6. The method of claim 1 , wherein the electrostatic interactions are reduced by administering about 50 mmol/L to about 500 mmol/L of CaCl 2 to the bodily sample.
7. The method of claim 2 , the detectable label being used to quantify the amount of β-defensin in the sample.
8. The method of claim 1 , the bodily sample comprising a bodily fluid.
9. The method of claim 8 , the bodily fluid selected from the group consisting of saliva, genital secretions, cervicovaginal lavage, bronchoalveolar lavage, blister fluid, and combinations thereof.
10. A method of quantifying β-defensin in a bodily sample of a subject comprising:
obtaining a bodily sample from the subject;
reducing the electrostatic interaction between β-defensin and negatively charged moieties in the bodily sample;
contacting the bodily sample with a capture antibody or an epitope binding fragment thereof, the capture antibody or an epitope binding fragment thereof being immunoreactive with the β-defensin;
contacting the bound β-defensin with a detection antibody or epitope binding fragment thereof, the detection antibody or epitope binding fragment thereof being coupled to a detectable label;
detecting the detectable label in the sample; and
correlating the amount of detectable label in the sample to an amount of β-defensin in the bodily sample.
11. The method of claim 10 , the step of reducing the electrostatic interaction between β-defensin and negatively charged moieties in the sample comprising adding positively charged moieties to the bodily sample at an amount effective to reduce the electrostatic interactions between β-defensin and negatively charged moieties in the bodily sample.
12. The method of claim 11 , the positively charged moieties comprising cations.
13. The method of claim 12 , the cations being selected from the group consisting of Mg 2+ , Ca 2+ , and combinations thereof.
14. The method of claim 10 , wherein the electrostatic interactions are reduced by administering about 50 mmol/L to about 500 mmol/L of CaCl 2 to the bodily sample.
15. The method of claim 10 , the bodily sample comprising a bodily fluid.
16. The method of claim 15 , the bodily fluid selected from the group consisting of saliva, genital secretions, cervicovaginal lavage, bronchoalveolar lavage, blister fluid, and combinations thereof.
17. A method of quantifying β-defensin in a bodily sample from a subject comprising:
obtaining a bodily sample from the subject;
adding CaCl 2 to the bodily sample at an amount effective to reduce electrostatic interaction between β-defensin and negatively charged moieties in the bodily sample
contacting the bodily sample with a capture antibody or an epitope binding fragment thereof, the capture antibody or an epitope binding fragment thereof being immunoreactive with the β-defensin;
contacting the bound β-defensin with a detection antibody or epitope binding fragment thereof, the detection antibody or epitope binding fragment thereof being coupled to a detectable label;
detecting the detectable label in the bodily sample; and
correlating the amount of detectable label in the sample to an amount of β-defensin in the bodily sample.
18. The method of claim 17 , the detection antibody or epitope binding fragment thereof being biotinylated.
19. The method of claim 17 , the bodily sample comprising bodily fluid.
20. The method of claim 19 , the bodily fluid selected from the group consisting of saliva, genital secretions, cervicovaginal lavage, bronchoalveolar lavage, blister fluid, and combinations thereof.