New recommendations from EULAR on interpreting the results of an- tinuclear antibody (ANA) testing advised taking the test methodology into account because of differences inperformance.
ANA results vary not only by the testbeing used but also by the underlyingdisease they are being used to assess,warned Prof. Pier Luigi Meroni, director ofthe Immunorheumatology Research Laboratory at the IRCCS Istituto AuxologicoItaliano in Milan.
“Antinuclear antibody testing is a knowndiagnostic tool. But the recent advancesin methodologies strongly suggest that wehave to update our knowledge for a betterinterpretation of the results,” Prof. Meronisaid in his presentation at the congress.
There is “no doubt that ANA testing isuseful,” he continued, adding that ANAis used as a primary screening tool inmany rheumatic diseases, notably systemic lupus erythematosus (SLE), primarySjögren’s syndrome, and systemic sclerosis. It’s also recently been suggested as animportant entry criterion for the classification of SLE.
In fact, the 2019 SLE classification
criteria – developed by EULAR in col-
laboration with the American College of
Rheumatology (ACR) – state that “testing
by immunofluorescence on HEp- 2 cells or a
solid-phase ANA screening immunoassay
with at least equivalent performance is
highly recommended,” Prof. Meroni said.
The ideas underpinning that recommendation was that “ANA expression isinvariable in SLE, and that ANA-negativelupus is quite rare,” he explained. Also, asSLE expression persists over time, ANAtesting could be used for classification atany point in the disease course. These assumptions have been borne out in severalstudies, with very small percentages ofpatients (6% or less) having ANA-negative lupus, and more than 80% having apositive HEp- 2 test over time, even withimmunosuppressive treatment.
Which test methodology to use?
There are several methods that can beused to detect ANA, including the preferred HEp- 2 indirect fluorescence assay(IFA), several solid-phase assays (SpA),and line- or dot-blot immunoassays. Theissue is which assay should be used inwhich disease?
The performance of a particular assaycan depend on the disease in which theyare used. For instance, while the HEp- 2 IFAand SpA are equivalent in SLE and in otherconnective tissue diseases, “this is notthe case for other autoimmune diseases inwhich basically we don’t know exactly allthe autoantigens,” Prof. Meroni explained.“Most of the autoantigens are undefined.
They cannot be found in solid-phase kits,
Importantly, neither the IFA nor the SpA
is superior to the other. “We just say that
one technique can detect relevant antibod-
ies that are not detectable by the other
one, and maybe the combination of the
two techniques can be the right strategy
to get the highest sensitivity,” Prof. Meroni
“Clinicians should be aware of the type
of assay used for ANA detection,” he said,
“because there are strong differences in
the performance, for example between IFA
and SpA, and such differences can have
important clinical and relevant
The test selected will depend on if the
aim is to exclude or confirm a disease, and
the optimal strategy will depend on pre-
test probability. For instance, IFA is more
sensitive than SpA for SLE and scleroder-
ma, whereas IFA is less sensitive than SpA
for Sjögren’s. For SLE, it is suggested to
use both the IFA and SpA. A combination
of both tests is also considered optimal for
scleroderma. SpA testing offers the best
sensitivity for Sjögren’s.
“The story is a little bit more complicated for inflammatory myopathies in whichwe don’t have assays able to detect allthe autoantibodies,” Prof. Meroni said. Inthat situation, several different techniqueshave to be used to check if the SpA resultsfit with the IFA pattern.
In 2019, the ACR released its own
position statement on ANA testing, high-
lighting that it supported the use of the
HEp- 2 IFA assay as the preferred option
for ANA testing and that labs should
specify the methods being used to test
for ANA when reporting their results. The
ACR position statement also noted that
“ordering healthcare professionals should
select specific ANA subserologies based
on a patient’s signs and symptoms and
when there is a high pretest suspicion for
a specific condition.”
Prof. Meroni disclosed serving as a con-
sultant to AbbVie, Inova Diagnostics, Mer-
ck Sharp & Dohme, Pfizer, Thermo Fisher
Scientific, and UCB.
Antinuclear antibody test interpretation
guidance gets refreshed