Statistics for autoimmune disorders are daunting. According to the National Institutes of Health (NIH), up to 23.5 million Americans suffer from more than 80 different autoimmune conditions, which are chronic and can be life-threatening. The disease is one of the top 10 leading causes of death in girls and women up to 64 years of age, and the NIH estimates that annual direct health care costs are in the range of $100 billion.

Contrast these numbers with comparable statistics for cancer: That disease strikes up to nine million Americans, and annual cancer costs are estimated at $57 billion, according to the NIH.

While diagnosis of autoimmune disorders—which run the gamut from rheumatoid arthritis to vasculitis to antiphospholipid syndrome (APS)—has become more accurate, the diseases are still notoriously elusive and difficult to determine. With no straightforward method to diagnose patients, clinicians rely on a combination of methods, including blood tests to measure auto antibodies, inflammation and organ function, and clinical symptoms. One of the primary autoimmune tests—Antinuclear Antibody (ANA)—is not standardized, even after decades of use.

Standardization Remains an Issue

“Standardization has always been a major topic, particularly for cardiolipin testing,” says Nina Olschowka, product manager of central marketing at Phadia, based in Germany. “The issue is that reproducibility in different labs and with different tests is limited.”

Michelle Altrich, director of new test development for IBT Laboratories, Lenexa, Kan, says, “For many autoimmune diseases, there is no gold standard for diagnosing. Markers have been identified for a lot of autoimmune diseases, but the connection between the marker and the diseases is unclear. We don’t know if the marker caused the disease, because it could be present without the disease.”

For many years, the technology of choice for autoimmune testing has been Immunofluorescent Antibody Analysis (IFA)—a sensitive, but subjective, largely manual method that relies on qualified and trained individuals to do the analysis.

According to Silvana Costa Manning of Euroimmun US, Bronton Township, NJ, IFA testing presents a particularly thorny problem for labs—which are under the gun to deliver quality test results quickly while continually finding ways to decrease costs (a necessity in this era to improve patient care), respond to stringent insurance regulations, and maintain a competitive position in the lab.

“To maximize efficiency, assays are automated or split out among a number of benches and shifts,” its managing director says. “This may diminish the technician’s ability to see the whole picture in determining a final test result.”

According to Manning, IFA testing for the most part has not kept pace with lab management’s streamlining efforts to produce a number of results with a minimum of labor.

“In general, IFA is stimulating, challenging, and requires low capital investment and provides a vast amount of information,” Manning says. “However, in a number of instances, multiple substrates must be tested and read to reach a final lab result.”

This is where IFA falls short. The available IFA technology does not fall in step with the lab, where one step or multiplex capability and speed are essential.

“At Euroimmun, our premise is that the ideal environment for the lab technician would offer the ability to see the full picture, raising the level of available information when making a determination,” Manning says. “The fact that this ideal may be achieved while speeding up the availability of results is an added bonus.”

Euroimmun has met its goal with the company’s proprietary IFA Biochip Mosaic System, which provides IFA multiplexing. Available for large-, medium-, and small-volume labs, it has made IFA testing—specifically Antineutrophilic Cytoplasmic Antibody (ANCA, a family of auto antibodies that indicates vasculitus) testing—easier and more comprehensive. Turnaround time is significantly improved without additional cost. Previously, ANCA testing required multiple assays, often performed at different times.

The ANCA Granulocyte Mosaic 23 System contains six biochips in one well, making one-step testing a reality. Two biochips are coated with human granulocytes (a type of white blood cell); one is treated with ethanol and the other with formalin, and utilized in the initial reading of c-ANCA and p-ANCA. The remaining four biochips are the control chips used to assist in the reading: HEP2, primate liver, PR3 antigen, and MPO antigen.

In one well, the lab can now simultaneously perform ANCA testing for c-ANCA and p-ANCA. In the same well are all the control biochips that allow technicians to read the granulocytic patterns, and results are available in 2 ½ hours.

“The need to check another slide, to look up other test results, delay test results awaiting a Hep2 run, etc, is rapidly becoming a thing of the past,” Manning says. “Multiplexed ANCA testing is a streamlined, coherent approach with excellent turnaround time and cost management.”

This same approach may be used when performing many other IFA test panels, such as Celiac testing, dermatological antibody testing, neuronal antibody, and mitochondrial antibody testing.

Testing for Multiple Parameters

GenBio, San Diego, offers an ELISA diagnostic for such autoimmune diseases as systemic lupus erythematosus that allows individual patients to be tested for multiple parameters simultaneously. According to its president and CEO, Fred Adler, ImmunoDOT is an easy-to-use, visually interpreted, rapid-test-format, dipstick ELISA. Up to five or six different tests may be completed simultaneously on a single patient specimen, making it ideal for labs that require fast, flexible, and reliable diagnostic tests in a single patient format.

ImmunoDot

A sophisticated test format, ImmunoDOT is a multiparameter, nitrocellulose ELISA with sensitivities and specificities similar to reference-method western blots, and equals or surpasses traditional ELISA formats for sensitivity and specificity, Adler says. It is used in labs preferring rapid turnaround, rather than batching, in medium and small hospitals, clinics, and group practice office labs.

The only expense for ImmunoDOT equipment is the modestly priced dry bath workstation designed to hold the disposable reaction vessels included with the kits.

The ImmunoDOT Autoimmunity Screening Panels detect antinuclear antibodies for specific diagnostically significant nuclear antigens and measure individually antibodies to the most common nuclear antigens: deoxyribonucleic acid, Sjogren’s syndrome antigen A and antigen B, ribonucleoprotein, Smith antigen, Scl-70 antigen, and Jo-1 antigen.

The ImmunoWELL DNA Antibody Test is an enzyme immunoassay (EIA) for the quantitative detection of antibodies to double-stranded DNA in serum and is used as an aid in the diagnosis of autoimmune diseases.

According to Adler, APS lab testing has greatly improved during the past 2 decades. Kits designed before the 1990s did not reliably detect anti-beta-2 glycoprotein, the most significant disease-specific component. In addition, many normal, healthy subjects have various anticardiolipin reactivities, and these false reactions minimized this assay’s clinical usefulness.

During the 1990s the complex interaction of aCL and a fatty acid, phosphotydl serine, was defined and shown to be a specific and diagnostic antibody response. GenBio’s Cardiolipin is standardized to ensure a reproducible and sensitive measure of this anti-beta-2 glycoprotein reaction. An additional beta-2 glycoprotein test is not required.

The ImmunoWELL Cardiolipin Antibody (IgG) Test and the ImmunoWELL Cardiolipin Antibody (IgM) Test are EIA assays that measure specific IgG and/or IgM aCL in human serum. The ImmunoWELL test provides highly reproducible results expressed in units that are standardized against an internationally recognized reference preparation. Levels of aCL antibodies are reported as GPL or MPL units.

Automated Options

Phadia offers the unique EliA system, which is specifically designed to offer clinically relevant results in autoimmunity testing while meeting the laboratory demands of efficient and cost-effective workflow. The ImmunoCAP 250 system is highly automated, requiring minimal hands-on time, and processes samples from test request to final result with minimal intervention.

Bar code identification of all reagents, mainframe worklist downloads, or bar code/manual sample identification combine with onboard dilution to reduce manual handling and minimize errors. Overnight runs with automatic shutdown after the last sample and an automated wake-up give the option of increasing the instrument’s available working hours.

The modular system allows for patient-specific, economical routine use, with calibration required only monthly. Wide measuring ranges and reproducible results optimize throughput and reduce the secondary-level testing required. Results can be turned around the same day.

“Our instruments are unique as no other specialized autoimmune company has instruments this size,” Olschowka says. “The ImmunoCAP 250 can run about 350 tests a day, plus overnight runs.”

The instrument had a large panel of clinically relevant autoimmune markers, some of which are known to be the best in the market, Olschowka says. These include the EliA Celikey test for antibodies against tissue transglutaminase, a diagnostic marker for celiac disease; EiA CCP, an important marker in rheumatoid arthritis; and ANAs.

IBT Laboratories markets an index test for chronic urticaria, a common skin disorder affecting up to 1% of the general population and characterized by chronic hives present for at least 6 weeks.

Based on physician needs, the company started developing the test in 2006 as a lab-developed test offered in its CLIA-certified labs. The test is not FDA-cleared as a kit.

“Our sales have been increasing since that time,” Chief Business Officer Maureen Loftus says. “We are in the high double digits every year.”

According to Altrich, the product is unique because it is a functional assay that uses donor basel cells as a reagent control that is incubated with patient serum. The two-stage test takes 2 days, and the sample serum can be shipped overnight at ambient temperature. After the ex vivo incubation, the cells are centrifuged, and the supernatant is recovered for histamine analysis. Using a quantitative enzyme immunoassay, the histamine is measured and compared to the total histamine in the base.

While the gold standard for autoimmune testing remains elusive, these companies and others like them are moving forward in the quest for accurate and quick diagnosis of these baffling conditions.

Shannon Rose is a freelance health and medical writer based in Temecula, Calif.