ANA Masterclass Flashcards
What structures and autoantigens are most frequently targeted by ANA (antibodies to nuclear antigens)?
What are the steps involved in ANA screening?
First step (see also Solomon et al., 2002):
The gold standard for screening for ANA is the indirect immunofluorescence (IIF) assay. This assay is sensitive but its specificity depends on the chosen cut-off. Substrate for the IIF are Hep‐2 cells (Hep2), a human larynx epithelioma cell line, or primate liver. Result of IIF – the antibody-antigen-reaction – is a special pattern of immunofluorescence, depending on the antigen and its localisation and role in the nucleus. IIF is the best screening method and should always be done first, but if positive, more specific testing is needed. In an international consensus (https://www.anapatterns.org/index.php, Damoiseaux et al., 2019), 29 so called AC (anti cell)-patterns have been defined. Another possibility to perform ANA-screening is a collective enzyme-linked immunosorbent assay (ELISA) which directly tests for many nuclear antigens simultaneously. Also here, if the signal is positive, more specific ELISAs are required to define the target antigen.
Second step
Depending on the test result (the IIF-pattern) and information about the patient’s symptoms the suspected autoantibodies can be more specifically detected, which is usually done by ELISA or protein blot; in this case only the reactivity to one antigen is tested with each ELISA. So far, ELISA-kits are commercially available for most but not all autoantigens, and some may only be performed by selected reference laboratories. In most cases, laboratories offer a screen, i.e. they test the most frequent ANA reactivities using the ELISA or the protein blot method. It would be just as wise to test only individual antigens, depending on the pattern. Depending on the sample and other clinical, instrumental and laboratory findings, antigen diagnostics can then be expanded. Not in all cases can the target antigen be correctly predicted by the pattern.
Furthermore, in IIF also antibodies against parts of the cytoplasm of HEp2 cells can be detected, even if they are not so easy to determine with IIF. The association of only few of them to specific autoimmune diseases could be found so far, e.g. antibodies against tRNA synthetases (Jo-1, PL-7, PL-12: antisynthetase syndrome) or antibodies against mitochondria (primary biliary cholangitis (PBC)). The clinical significance of many other of cytoplasmatic autoantibodies could not be clarified so far.
What database can you refer to for ANA pattern classification?
ICAP - International Consensus on ANA Patterns (https://www.anapatterns.org/index.php) - 29 different ANA patterns (classified into 3 groups - nuclear, cytoplasmic and mitotic). Each pattern and sub-pattern is assigned an anti-cell (AC) pattern from AC-1 to AC-29.
What clues on a general ‘systems review’ might indicate possible CTD and be of significance in the context of a positive ANA?
What diagnostic findings may potentially be related to CTD?
What diagnostics should be considered in the case of a positive ANA titre (depending on the pattern/specific ANA) and the overall context?
What are some recommended laboratory diagnostics in case of positive ANA/which could be related to connective tissue disease?
What is the prevalence of a SLE vs. the prevalence of a positive ANA test?
Almost all patients with SLE are ANA-positive – but lupus is a rare disease with a prevalence of about 0.05% in Europe.
On the other hand, in about 3 % of the population, ANA titres of 1:320 can be detected, or even more than 10% have a titre of 1:80. Both findings together make it clear: The probability that a person with a positive ANA titre suffers from lupus is very low. The vast majority of individuals with positive ANA titres is not affected by any rheumatological diseases.
What is the significance of a dense fine speckled (DFS70) antibody?
There is one type of nuclear antibodies that is not associated with connective tissue diseases: the DFS70 antibodies (dense fine speckled, 70 kD) can be responsible for a positive ANA test result. Even if it should not be considered an exclusion criterion, in case of positive DFS70 fluorescence pattern (and negative findings of other specific ANA), it can be assumed with higher probability that the patient is healthy from a rheumatological point of view (Mariz et al., 2011).
What is the frequency of elevated ANA titres in SLE?
95-100%
What is the frequency of elevated ANA titre in cutaneous lupus?
20-60%
What is the frequency of elevated ANA titre in drug-induced lupus?
95-100%
What is the frequency of elevated ANA titre in MCTD?
100%
What is the frequency of elevated ANA titre in systemic sclerosis?
60-80%
What is the frequency of elevated ANA titre in Sjogren’s syndrome?
40-95%
What is the frequency of elevated ANA titre in dermatomyositis?
30-80%
What is the frequency of elevated ANA titre in RA?
30-50%
What is the frequency of elevated ANA titre in sarcoidosis?
5-30%
What is the frequency of elevated ANA titre in PAN?
20%
What is the frequency of elevated ANA titre in JIA?
25-65%
What is the frequency of elevated ANA titre in Hashimoto’s thyroiditis?
45%
What is the frequency of elevated ANA titre in Grave’s disease?
50%
What is the frequency of elevated ANA titre in autoimmune hepatitis?
75%
What is the frequency of elevated ANA titre in Primary Biliary Cholangitis (PBC)?
80-100%
What is the frequency of elevated ANA titre in Hepatitis C?
5-54%
What is the frequency of elevated ANA titre in pulmonary fibrosis?
20-60%
What is the frequency of elevated ANA in ulcerative colitis?
25%
What viral infection can cause a positive ANA result?
EBV infection (B-cell activation)
What is the frequency of elevated ANA titre in the healthy population?
The frequency of elevated ANA titre (>/= 1:80) in the healthy population is 5-18% (depending on the age and height of the titre).
1:40 –> 32%
1:80 –> 13%
1:160 –> 5%
1:320 –> 3.2%
