Portrait of the HEp2 cell, the pet of immunofluorescence professionals


HEp2 cells -- centromere B

Anti-Centromere B on HEp2 cells

HEp2 cells are held dear in autoimmune diagnostics. They are invaluable for people engaged in analysing autoantibodies, as E. coli is for molecular biologists or mice for toxicologists.

In spite of a wide range of other suitable methods and technologies, determination of autoantibodies with indirect immuno-fluorescence assays (IFA) on human epithelioma (HEp2) cells still contributes significantly to the diagnosis of autoimmune diseases. The widely recognised advantages of this method are high sensitivity and a broad spectrum of antibodies that can be analysed simultaneously. In addition to mere detection of antibodies a characteristic fluorescence pattern and staining of metaphase and cytoplasmic cells offer supplementary information.

When an autoimmune disease is suspected, the HEp-2 test usually is the first line test. Any positive result is then followed up by a step-wise diagnostic approach, including other immunological tests like ELISA (enzyme-linked immunosorbent assay) for single antibody specificities or immunoblot tests. Continue reading

Liver Disease Diagnostics: Antibody-based Diagnosis of Autoimmune Liver Diseases


Liver Disease Diagnostics: Antibody-Based Diagnosis of Autoimmune Liver Disease

For the launch of our Liver-9-Line immunoblot test (to our press release “Liver Disease Diagnostics by Immunoblot” of May 16, 2011), I dug through a pile of literature on the topic of autoantibody-based diagnosis of autoimmune diseases of the liver. In the last week I picked it all up again and worked through it systematically.

This picture shows the interiour surface of the liver. It's a reproduction of a lithograph plate from Gray's Anatomy.

The interior surface of the liver. A reproduction of a lithograph plate from Gray’s Anatomy.

The reason for my renewed interest is that we brought four more ELISA tests for liver diagnostics to the market two weeks ago. They are the Anti-LKM-1, Anti-SLA, Anti-gp210, and Anti-Sp100 tests, all designed for fully automated autoimmune diagnosis with our Alegria system. All four test systems assist the formulation of a diagnosis when autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are suspected, or for differential diagnosis when another disorder of the liver is assumed. Continue reading

A short History of Indirect Immunofluorescence Technology


The GRÜNER CLUB AUTOIMMUN blog featured a fine post about the history of indirect immunofluorescence. In that article my Austrian colleague Barbara Fabian, community manager of GRÜNER CLUB AUTOIMMUN, described in great detail how indirect immunofluorescence technology, or: IFT, and also referred to as IIF assay,  has become an indispensible tool of autoimmune disease diagnostics over the last two decades, and how IFT has become a standard laboratory technique used in serological autoimmune diagnostics.

Without further ado I have translated Barbara’s post in order to make you this text, and especially the interesting images, available. – Here it is:

The Development of Indirect Immunofluorescence Technology (IFT)

by Barbara Fabian, MSc, Community Manager of GRÜNER CLUB AUTOIMMUN

Over the last 20 years, the detection of autoantibodies has developed into an indispensible component of autoimmune diagnostics. Along with serological and clinical data, autoimmune status has become an important building block in the formation of diagnoses. Continue reading

Rheumatology Diagnostics: Indirect Immunofluorescence Tests (IIF assays) on HEp-2 Cells


Immunofluorescence patterns help eliminate “false positives” in diagnosing autoimmune rheumatic diseases

The detection of anti-nuclear antibodies, the ANA test, is a clear (laboratory-) diagnostic indicator of rheumatic autoimmune disease. One of the standard laboratory tests for the detection of these antinuclear antibodies is IIF, the indirect immunofluorescence assay, on human HEp-2 cells (ANA-HEp-2 test).

This pictures shows anti-RNP pattern on HEp-2 cells after ANA-HEp-2 indirect immunofluorescence assay

ANA-HEp-2 indirect immunofluorescence test (IIF): antibodies against RNP (ribonucleoproteins) – interphase nucleoli: coarse granular positive, nucleoli neglected; mitotic cells: negative (400x) – © ORGENTEC Diagnostika, Mainz

However, for up to 13% of healthy individuals, indirect immunofluorescence may detect anti-nuclear antibodies. Most of these healthy people will not develop an autoimmune disease – despite the positive ANA test. It is thus a challenge for the physician to differentiate these healthy, false-positive patients from those ANA-positive patients who already have an inflammatory rheumatic disease or who truly have an increased risk of developing such an autoimmune disease.

 

Several very specific IIF patterns

In a large study, Brazilian IIF experts have now worked out the fundamental differences between the ANA-HEp-2 test results on serum samples from healthy individuals and the immunofluorescence patterns from serum samples of patients with rheumatic disease; they have described various IIF patterns that can be used to differentiate between the two patient groups (Mariz et al. 2011). This study was published a few weeks ago in the January issue of Arthritis & Rheumatism, the journal of the American College of Rheumatology (ACR). In their article, the scientists from the Universidade Federal de São Paulo, Brazil, explain in detail that there are several very specific immunofluorescence patterns in the ANA-HEp-2 assay with which the autoimmune rheumatic diseases (ARD) are truly associated. Continue reading

Autoantibodies precede manifestation of Lupus by years


Antibodies to various autoantigens may be present in sera of patients who will develop Lupus erythematosus up to seven years before onset of disease symptoms

 

Immunoflurescence image of anti-dsDNA-antibodies for the diagnosis of SLE

Autoantibodies against dsDNA are diagnostic markers for Systemic Lupus Erythematosus.

Autoantibodies are specific and sensitive biomarkers for autoimmune diseases and indispensible diagnostic tools. They may also be involved in pathogenic processes underlying the disease and will potentially occur in sera of apparently healthy people long before onset of the first symptoms.

C. Eriksson, S. Raantapaa-Dalquist and their colleagues from Umeå University in Sweden have focused on this preclinical phase in the development of Systemic Lupus Erythematosus (SLE). Continue reading