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Further Characterization of PBC Related Autoantigens: The Role of Nucleoporin p62 (Nup62)

Primary biliary cirrhosis (PBC) is an immune-mediated chronic cholestatic disease characterized by specific anti-mitochondrial antibody responses directed against members of the 2 oxo-acid dehydrogenase complexes (M2 antigen) and in particular the inner lipoyl domains of the E2 subunits of pyruvate and other oxoacid dehydrogenase complexes. These strong diagnostic requirements have been met by our strategy to establish an M2 target by combination of full-length human recombinant PDC-E2, BCOADC-E2, and OGDC-E2 antigens (c.f. DIARECT newsletter 2, 2006).

Additional useful diagnostic markers for PBC are antinuclear antibodies (ANA), and, therefore, during the last 2 ½ years we have expanded our toolbox of PBC-related diagnostic targets by adding recombinant human Sp100 and gp210 (newsletters 1 & 2, 2008).

The disease specificity of detection of gp210 auto-antibodies is close to 100%, but in contrast to M2 antibodies they are said to have profound diagnostic significance and may be closer associated with more active and severe disease and therefore poor prognosis.

Whereas the Sp100 autoantigen represents the target with highest coherency to multiple nuclear dot immunofluorescence pattern (MND IIF), autoantibodies reactive with the nuclear membrane glycoprotein gp210 appear to correlate with just one antigenic structure responsible for positive nuclear membrane IIF (Wesierska-Gadek et al. 2007). In addition to anti-gp210 reactivity, these authors described another strong reaction of PBC sera with a protein of approx. 60 kDa, which turned out to be a constituent of the nuclear pore complex (NPC), namely nucleoporin p62 (Nup62). There is now evidence that anti-Nup62 antibodies occur even more frequently than the autoantibodies against gp210 glycoprotein.

DIARECT's biotech facilities have recently established a full-length recombinant human Nup62 autoantigen, which has now been evaluated by clinically defined PBC sera (Prof. Dr. R. Klein, Tübingen University, Germany). Interestingly, several patient sera negative for M2 antibodies, but with a positive IIF pattern for nuclear membrane could be shown to be anti-Nup62 positive (figure).

Considering the increasing clinical importance of ANAs in PBC diagnostics, the prognostic value of all these antibodies (MND, NM, NPC) and their correlation with severity and progression of the disease is of great interest. With the availability of the major nuclear antigenic elements (Sp100, gp210, Nup62) as well as the classical M2 targets (PDC-, BCOADC-, OGDC-E2) as recombinant human autoantigens, their significance may now be determined.

DIARECT now offers the most complete panel of recombinant human targets for primary biliary cirrhosis autoantibody testing:

IVD of Celiac Disease - Usefulness of Deamidated Gliadin Targets

Celiac disease is a chronic intestinal disorder most probably caused by an abnormal immune reaction to wheat gliadin and related gluten components from barley and rye. Although the disease starts as intolerance to gliadins, antibodies to tissue-type transglutaminase (tTG) in the gut epithelium are also characteristic of the disease.

Historically, serologic tests for gliadin antibodies generally lacked the accuracy required for proper diagnosis due to missing deamidated epitopes within the authentic gliadin fraction formerly used in diagnostic test kits. Because the recognition of gliadin by intestinal T cells of genetically predisposed individuals is dependent on deamidation, a modification known to be promoted in the acidic environment during pepsin digestion. However, deamidation of gliadin is most probably mediated in vivo by tTG in a specific fashion. This modification introduces negative charges into gliadin (glutamate residues) and consequently increases the binding affinity for DQ2 and/orDQ8 type HLA class II molecules (Arentz-Hansen et al. 2000).

In comparison with native, nonmodified gliadin preparations, the diagnostic applications of deamidated gliadin synthetic (nona)peptides led to a considerable enhancement of diagnostic accuracy (Schwertz et al. 2004).

Since obviously no single test encompasses almost 100% specificity and sensitivity at the same time (Marietta et al. 2009) the current, recommended strategy for celiac disease diagnosis comprises a combination of IgA and IgG testing of anti-deamidated gliadin reactivity complemented by additional anti-IgA-tTG testing.

As a protein-focused biotech enterprise, DIARECT has successfully completed the recombinant protein approach by designing and producing a recombinant, deamidated γ-gliadin antigen.

Proof of the superior quality of SurModics Stabilizer StabilGuard in Luminex assays

A brand-new article, published in Clinical and Vaccine Immunology focuses on the elimination of  false-positive reactivity of patient sera in a Luminex assay for pneumococcal antibodies.  In this study SurModics stabilizer StabilGuard was proven to completely eliminate nonspecific binding to uncoupled MicroPlex microspheres by resuspending the uncoupled microspheres in StabilGuard.

Pickering, J.W., Larson, M.T., Martins, T.B., Copple, S.S., Hill, H.R. (2010) Elimination of False-Positive Results in a Luminex Assay for Pneumococcal Antibodies. Clinical and Vaccine Immunology 17: 185-189

Calendar 2010 -  Meet DIARECT!

Meet the DIARECT team at the following events:


Analytica 2010
Date:        March 23 – 26, 2010  
Location: Munich, Germany

The 7th International Congress on Autoimmunity
Date:        May 5 – 9, 2010  
Location: Ljubljana, Slovenia

12. Autoimmuntage in Bernau
Date:        May 20 – 21, 2010  
Location: Bernau, Germany

BIO JAPAN 2010
Date:        June 30 – July 2, 2010  
Location: Tokyo, Japan

AACC Annual Meeting 2010
Date:        July 25 – 29, 2010  
Location: Anaheim, California

AMLI 23rd Annual Meeting
Date:        August 7 – 10, 2010  
Location: Orlando, Florida

7. Jahrestagung DGKL
Date:        September 29 - October 02, 2010  
Location: Mannheim, Germany

Biotechnica 2010
Date:        October 5 – 7, 2010  
Location: Hannover, Germany

ACR / ARHP 2010 Annual Scientific Meeting
Date:        November 6 – 11, 2010  
Location: Atlanta, Georgia

Medica 2010
Date:        November 17 – 20, 2010  
Location: Düsseldorf, Germany

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