Inflammatory bowel disease and irritable bowel syndrome IBD-first chek diagnostic panel
A method for the differentiation of inflammatory bowel disease (IBD) from irritable bowel disease (IBS) followed by distinguishing ulcerative colitis and Crohn's disease from other gastrointestinal illnesses. This highly differential method first uses the presence of elevated lactoferrin as a marker of intestinal inflammation to differentiate IBD from IBS. Patients suspected of IBD are then analyzed for fecal anti- Saccharomyces cerevisiae antibodies (ASCA) as an indicator of Crohn's disease and fecal anti-neutrophil cytoplasmic antibodies (ANCA) as an indicator of ulcerative colitis. IBD patients are further monitored for intestinal inflammation using fecal lactoferrin to evaluate the effectiveness of medical therapy and to predict relapse. The apparatus comprises either a qualitative enzyme-linked immunoassay or other immunoassay that utilizes antibodies specific to human immunoglobins for the measurement of total endogenous lactoferrin, ASCA and ANCA in human feces. The method and apparatus can be used by healthcare providers to identify IBD and distinguish ulcerative colitis from Crohn's disease.
1. A method for testing fecal samples from persons for diagnosis, the method comprising:
obtaining a fecal sample from a person presenting with symptoms common to inflammatory bowel disease and irritable bowel syndrome;
diluting the sample;
determining that the sample contains an elevated level of lactoferrin compared to a lactoferrin level in a healthy control;
measuring the sample for an elevated level of anti- Saccharomyces cerevisiae antibodies (ASCA);
measuring the sample for an elevated level of anti-neutrophil cytoplasmic antibodies (ANCA);
upon determining that the sample contains an elevated level of anti- Saccharomyces cerevisiae antibodies compared to an anti- Saccharomyces cerevisiae antibody level in a healthy control and not an elevated level of anti-neutrophil cytoplasmic antibodies, diagnosing the person with Crohn's disease; and
upon determining that the sample contains an elevated level of anti-neutrophil cytoplasmic antibodies compared to an anti-neutrophil cytoplasmic antibody level in a healthy control and not an elevated level of anti- Saccharomyces cerevisiae antibodies, diagnosing the person with ulcerative colitis.
2. The method of claim 1 , wherein a diagnosis of inflammatory bowel disease may be concluded based upon the sample containing an elevated level of lactoferrin.
3. The method of claim 1 , wherein the lactoferrin, anti- Saccharomyces cerevisiae antibodies and anti-neutrophil cytoplasmic antibodies are measured using one of enzyme-linked immunoassays, lateral flow membrane tests and immunoassays utilizing antibodies.
4. The method of claim 1 , wherein determining that the sample contains an elevated level of lactoferrin is based on a qualitative ELISA.
5. The method of claim 1 , wherein determining that the sample contains an elevated level of lactoferrin is based on a quantitative measurement.
6. The method of claim 1 , further comprising:
contacting the sample with immobilized polyclonal antibodies to endogenous lactoferrin to create a treated sample;
contacting said treated sample with enzyme-linked polyclonal antibodies such that the enzyme-linked polyclonal antibodies are allowed to bind to captured endogenous lactoferrin creating an enzyme-linked antibody bound sample;
adding a substrate to the enzyme-linked antibody bound sample to create a readable sample; and
determining the optical density of said readable sample at 450 nm.
7. The method of claim 1 , further comprising:
contacting the sample with antigens of Saccharomyces cerevisiae to create a treated sample;
contacting the treated sample with polyvalent antibodies to human immunoglobulin conjugated to an enzyme such that the polyvalent antibodies are allowed to bind to capture anti- Saccharomyces cerevisiae antibodies creating an enzyme-linked antibody bound sample;
adding a substrate to the enzyme-linked antibody bound sample to create a readable sample; and
determining the optical density of the readable sample.
8. The method of claim 1 , further comprising:
contacting the sample with neutrophil cytoplasmic antigens to create a treated sample;
contacting the treated sample with polyvalent antibodies to human immunoglobulin such that the enzyme-linked polyvalent antibodies are allowed to bind to capture anti-neutrophil cytoplasmic antibodies creating an enzyme-linked antibody bound sample;
adding an enzyme substrate to the enzyme-linked antibody bound sample to create a readable sample; and
determining an optical density of the readable sample at 450 nm.
9. A method for testing fecal samples from persons for diagnosis, the method comprising:
obtaining a fecal sample from a person presenting with symptoms common to inflammatory bowel disease and irritable bowel syndrome;
diluting the sample;
contacting the diluted sample with immobilized polyclonal antibodies to endogenous lactoferrin to create a first treated sample;
contacting said first treated sample with enzyme-linked polyclonal antibodies such that the enzyme-linked polyclonal antibodies are allowed to bind to captured endogenous lactoferrin creating an enzyme-linked antibody bound sample;
adding a substrate to the enzyme-linked antibody bound sample to create a readable sample;
determining the optical density of said readable sample at 450 nm;
determining that the sample contains an elevated level of lactoferrin compared to a lactoferrin level in a healthy control;
measuring the sample for an elevated level of anti- Saccharomyces cerevisiae antibodies (ASCA);
measuring the sample for an elevated level of anti-neutrophil cytoplasmic antibodies (ANCA);
upon determining that the sample contains an elevated level of anti- Saccharomyces cerevisiae antibodies compared to an anti- Saccharomyces cerevisiae antibody level in a healthy control and not an elevated level of anti-neutrophil cytoplasmic antibodies, diagnosing the person with Crohn's disease; and
upon determining that the sample contains an elevated level of anti-neutrophil cytoplasmic antibodies compared to an anti-neutrophil cytoplasmic antibody level in a healthy control and not an elevated level of anti- Saccharomyces cerevisiae antibodies, diagnosing the person with ulcerative colitis.
10. The method of claim 9 , further comprising:
contacting the sample with antigens of Saccharomyces cerevisiae to create a second treated sample;
contacting the second treated sample with polyvalent antibodies to human immunoglobulin conjugated to an enzyme such that the polyvalent antibodies are allowed to bind to capture anti- Saccharomyces cerevisiae antibodies creating an enzyme-linked antibody bound sample;
adding a substrate to the enzyme-linked antibody bound sample to create a readable sample; and
determining the optical density of the readable sample.
11. A method for testing fecal samples from persons for diagnosis, the method comprising:
obtaining a fecal sample from a person presenting with symptoms common to inflammatory bowel disease and irritable bowel syndrome;
diluting the sample;
contacting the sample with neutrophil cytoplasmic antigens to create a treated sample;
contacting the treated sample with polyvalent antibodies to human immunoglobulin such that the enzyme-linked polyvalent antibodies are allowed to bind to capture anti-neutrophil cytoplasmic antibodies creating an enzyme-linked antibody bound sample;
adding an enzyme substrate to the enzyme-linked antibody bound sample to create a readable sample;
determining an optical density of the readable sample at 450 nm;
determining that the sample contains an elevated level of lactoferrin compared to a lactoferrin level in a healthy control;
measuring the sample for an elevated level of anti- Saccharomyces cerevisiae antibodies (ASCA);
measuring the sample for an elevated level of anti-neutrophil cytoplasmic antibodies (ANCA);
upon determining that the sample contains an elevated level of anti- Saccharomyces cerevisiae antibodies compared to an anti- Saccharomyces cerevisiae antibody level in a healthy control and not an elevated level of anti-neutrophil cytoplasmic antibodies, diagnosing the person with Crohn's disease; and
upon determining that the sample contains an elevated level of anti-neutrophil cytoplasmic antibodies compared to an anti-neutrophil antibody level in a healthy control and not an elevated level of anti- Saccharomyces cerevisiae antibodies, diagnosing the person with ulcerative colitis.