Antinuclear Antibody Test

Antinuclear antibodies (ANAs) have emerged as valuable diagnostic markers in the field of autoimmune connective tissue diseases. These antibodies, which bind to cellular components within the nucleus of cells, including proteins, DNA, RNA, and nucleic acid-protein complexes, play a crucial role in the identification and diagnosis of various autoimmune disorders such as lupus, scleroderma, juvenile arthritis, Sjögren's syndrome, and dermatomyositis.

Browse all ANA products

Immunity and Autoimmunity

Human physiology encompasses a multitude of defense mechanisms to counteract pathogens, among which humoral immunity plays a vital role. This intricate defense mechanism triggers the production of large glycoproteins called antibodies, prompted by an immune stimulus. The orchestration of this immune response involves various cells of the immune system, including lymphocytes such as T-cells and B-cells, as well as antigen-presenting cells. These cellular components collaborate to mount an immune defense upon detection of foreign proteins, generating specific antibodies that bind to these antigens.

Under normal physiological conditions, lymphocytes that recognize self-antigens, also known as autoantigens, undergo either programmed cell death (apoptosis) or lose their functionality. This phenomenon, known as self-tolerance, ensures that lymphocytes do not initiate an immune response against human cellular antigens. However, in certain instances, this intricate process can malfunction, leading to the production of antibodies targeting human antigens. This aberrant immune response has the potential to trigger autoimmune diseases, characterized by the immune system attacking its own cells and tissues.

Antinuclear Antibody Test

• Immunofluorescence Assay
  The fluorescent immunohistochemistry technique using mouse liver tissues or imprints is commonly used to detect serum ANAs. Positive reactions are observed as fluorescent staining of cell nuclei. Diluting patient serum allows for approximate quantitative tests, with a positive reaction at 1:80 dilution being significant for diagnosing SLE. Different fluorescent patterns indicate specific antibody types associated with SLE and other connective tissue diseases. For example, a homogeneous staining pattern may indicate the presence of anti-dsDNA antibodies. Researchers have explored using protozoa like Trypanosoma or blood flagellates (e.g., Crithidia luciliae) as substrates to detect anti-dsDNA antibodies, offering high specificity and sensitivity. These harmless protozoa possess pure dsDNA within their kinetoplasts, enabling clear fluorescence at the flagellum-associated kinetoplast end. This approach shows promise for wider applications.

Figure 1. Diagrammatic representation of common nuclear patterns observed under fluorescence microscopy.
(Source: Kumar, Y. et al., 2009)

• ELISA
  ELISA is another commonly employed method for ANA testing. It utilizes specific antigens coated onto a microplate, allowing for the quantification of ANAs in the patient's serum. ELISA offers advantages such as increased sensitivity and the ability to detect low ANA titers.
• Radioimmunoassay
  Common methods for detecting ANAs include the Farr method and the filtration method. The Farr method involves labeling DNA with isotopes and measuring the binding activity between ANAs and labeled DNA. A binding rate above 20% is considered positive. The filtration method uses a membrane filter to separate complexed ANAs from free DNA. The complexed antibodies are retained on the filter while free DNA passes through. These methods provide effective means for ANA detection in clinical settings.
• Immunoblotting
  In this method, mixed antigens are separated using gel electrophoresis and transferred to a nitrocellulose membrane. Acid or radioisotope-labeled antibodies are then used for detection and analysis. This approach allows for multiple analyses on the same solid phase without the need for antigen purification. It offers high sensitivity and specificity, making it widely used to detect various autoantibodies in the serum of patients with autoimmune diseases, including anti-Sm, anti-RNP, anti-SS-A, and anti-SS-B antibodies.

In addition, other methods such as flow cytometry, microarray, indirect hemagglutination, and complement fixation tests can also be used for ANA detection. The choice of these techniques depends on various factors, including the screening or specific ANA testing requirements, sensitivity and specificity of the test, availability, cost-effectiveness, and the time and expertise required to perform the test.

ANA-Related Antibody

ANA-Related Antigen

ANA-Related ELISA