Methods for detecting novel autoantibodies in Crohn's disease
Identification of autoantibodies associated with Crohn's disease useful in diagnosis and management using an innovative protein array technology, namely nucleic acid programmable protein arrays (NAPPA) and applications relating thereto. Overall, reactivity of IgG autoantibodies was stronger than that of IgA autoantibodies; however, IgA autoantibodies showed greater differential reactivity between cases and controls. Four IgA autoantibodies against SNRPB, PRPH, PTTG1 and SNAI1 were newly identified with sensitivities above 15% at 95% specificity, among which anti-SNRPB-IgA had the highest sensitivity of 24.0%. Autoantibodies associated with specific disease subtypes were also found.
1. A method for detecting an autoantibody of Crohn's disease in a subject comprising: disposing a plurality of DNA segments corresponding to different open reading frames of genes on an array, wherein each DNA segment comprises a nucleic acid sequence encoding a fusion tag; disposing a plurality of antibodies raised against the fusion tag on the array; translating the plurality of DNA segments to a plurality of polypeptides, wherein each polypeptide comprises the fusion tag and is captured by the antibodies raised against the fusion tag, wherein the plurality of captured polypeptides are selected from the group consisting of A4-Fla2, flaD, XAGE2, CDC42EP1, PPP1R13L, FOSL1, SNRPB, CCHCR1, LCORL, NUPL1, PRPH, PTTG1, SNAII and ORC2L, and incubating the array comprising the plurality of captured polypeptides with a sample collected from a subject suspecting having the disease, identifying the subject having Crohn's disease based, at least in part, on the presence of the polypeptide/autoantibody complex; and
administering a medical therapy selected from the group consisting of therapeutic steroids, immunomodulators, and biologics.
2. The method of claim 1 , wherein the disposing a plurality of DNA segments step further comprises coating the array with aminosilane.
3. The method of claim 1 , wherein the fusion tag is a C-terminal glutathione S-transferase (GST) fusion tag.
4. The method of claim 1 , wherein the disposing a plurality of antibodies step further comprises incubating the array in a printing buffer containing anti-GST polyclonal antibodies.
5. The method of claim 1 , wherein the translating step further comprises expressing polypeptides from the disposed plurality of DNA segments using a HeLa cell lysate-based in vitro transcription and translation (IVTT) protein expression system.
6. The method of claim 5 , wherein the expressing step further comprises incubating the array with a human IVTT mixture.
7. The method of claim 1 , wherein the plurality of captured polypeptides comprise A4-Fla2, FOSL1, PRPH, and XAGE2.
8. The method of claim 1 , wherein the plurality of captured polypeptides comprise A4-Fla2, FOSL1, PRPH, XAGE2, and SNRPB.
9. The method of claim 1 , wherein the plurality of captured polypeptides comprise A4-Fla2 and SNRPB.
10. The method of claim 1 , wherein the plurality of captured polypeptides comprise SNRPB.
11. The method of claim 1 , wherein the plurality of DNA segments corresponding to different open reading frames of genes are obtained from DNASU plasmid repository.
12. The method of claim 1 , wherein the plurality of DNA segments corresponding to different open reading frames of genes encode about 1900 different human proteins.
13. The method of claim 1 , wherein the sample is a blood or serum sample.
14. The method of claim 1 , wherein the disease is Crohn's disease.