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Imse Lab > ANA DETECTION > Flashcards

ANA DETECTION Flashcards

1
Q

Materials

A

⤳ SLE latex suspension
⤳ Black test cards
⤳ Mechanical rotator
⤳ Positive and Negative Controls
⤳ Disposable stirrers

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2
Q

Principle of the test

A

Passive Latex Agglutination

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3
Q

Meaning of ELISA

A

Enzyme-Linked Immunosorbent Assay

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4
Q

Meaning of CLIA

A

Chemiluminescence immunoassay

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5
Q

These assays can testfrom a broad range of antibodies if multiple nuclear antigens are coated onto a single test well, or for a specific
ANAs if each well is coated with a single antigen.

A

ELISA and CLIA

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6
Q

ELISA & CLIA depends on?

A

Depends on the type of nuclear antigen loaded on the well

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7
Q

Capable of detecting various diseases

A

Multiple nuclear antigens

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8
Q

Capable of detecting single or specific diseases

A

Specific nuclear antigen

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9
Q

These antigens use in commercial kits are derived from

A

Tissue extracts or produced by recombinant technology

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10
Q

It is used to screen for the presence of anas in addition to identifying specific anas

A

ELISA methods

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11
Q

There is a large variation in the performance of test produced by different manufacturers, which is influenced by the

A

Antigen preparation used

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12
Q

One study found sensitivities of elisa assays ranging from

A

69% to 98%

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13
Q

Specificities of elisa ranges from

A

81% to 98% when they were compared with IFF ANA method

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14
Q

Proportion of patients with a disease who test positive

A

Sensitivity

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15
Q

Proportion of patients without a disease who test negative

A

Specificity

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16
Q

If 100 patients known to have a disease were tested, and 43 test positive, then the test has 43% sensitivity. If 100 with no disease are tested and 96 return a completely negative result, then the test has 96% specificity.

A

True

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17
Q

Reagents

A

Ana reagent
Positive and negative control reagents
Wells

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18
Q

Qualitative method (screening) procedure

A
  1. Bring all reagents to room temperature and mix gently prior to use.
  2. Place the following on separate divisions of the same black test card.
  3. Place a drop of undiluted serum into the circles of the test slides
    ⤳ Patient’s Serum: 1 drop
    ⤳ Positive Control: 1 drop
    ⤳ Negative Control: 1 drop
  4. Add 1 drop of SLE Latex reagent to each sample on the black test card.
  5. Mix with the flat end of the pipette/mixer and spread fluid evenly covering the entire
    diameter of the test circle.
  6. Tilt the slide back and forth slowly for 3 minutes while observing for agglutination.
  7. Observe and Interpret results.
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19
Q

The test is considered as NEGATIVE when no difference in agglutination is observed between the specimen and negative control and POSITIVE sera must show distinct agglutination within _ minutes.

A

3

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20
Q

Agglutination indicates the level of ___ (especially anti-DNP) in the
range commonly found in the SLE.

A

antinuclear antibody

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21
Q

Findings in the sensitivity studies suggest that immunoassays may miss a significant proportion of ANA-positive patients and also yield a significant number of _____

A

false positive results.

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22
Q

Based on such studies, the ___ has recommended that the IIF test remains the gold standard for ANA testing and that clinical laboratories should specify the method they use when they are reporting the result.

A

ACR (American College of Rheumatology)

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23
Q

gold standard for ANA testing and that clinical laboratories should specify
the method they use when they are reporting the result.

A

Indirect Immunofluorescence Test
(IIF)

24
Q

FANA TESTING PATTERNS

A

HOMOGENOUS
SPECKLED
NUCLEOLAR
MIXED
CENTROMERE
PERIPHERAL

25
Q

CLINICAL SIGNIFICANCE

A

Screening method for SLE but not specific

26
Q

is a chronic systemic inflammatory disease that affects people usually at the peak age between 20 and 40 year

A

Systemic Lupus Erythematosus

27
Q

SLE is a chronic systemic inflammatory disease that affects people usually at the peak age between

A

20 and 40 years

28
Q

Women are much likely more to be affected than men by a ratio of ___.

A

9:1

29
Q

This autoimmune disease appears to originate from complex interaction between several factors, such as:

A

Environmental
Genetic susceptibility
Abnormalities within the immune system.

30
Q

⤳ In line with these, over ___ autoantibodies associated with SLE have been discovered.

A

100

31
Q

Autoantibodies includes

A

● double stranded DNA (dsDNA)
● histones, and other nuclear components, as well as
● autoantibodies to lymphocytes, erythrocytes, platelets, phospholipids, ribosomal components, and endothelium

32
Q

Laboratory diagnosis of SLE include

A

CBC, platelet count and urinalysis.

33
Q

Findings in lupus patients are

A

leukopenia and possible anemia and thrombocytopenia.

34
Q

Erythrocyte Sedimentation Rate (ESR) may be ___ even though
C-Reactive Protein tends to be low or normal.

A

Elevated

35
Q

When SLE is suspected, the first test typically done is a screening test for
Antinuclear Antibodies (ANA). These antibodies are present in most cases of SLE, antibodies that are
directed against antigens of

A

nuclei of mammalian cells.

36
Q

ANAs are present in over _____ of patients with active SLE and are used as a major marker for the disease.

A

95%

37
Q

ANAs are not specific only to SLE as it may be present in other
connective tissue diseases such as

A

Sjogren’s syndrome
Scleroderma
Polymyositis-dermatomyositis
RA.

38
Q

They can also be found in some individuals with other conditions, including:

A

Chronic infections
Cancer
Pregnancy

39
Q

Furthermore, up to ___ of healthy individuals and up to ___ of elderly people are ANA-positive

A

5%
30%

40
Q

are a heterogeneous group of antibodies that have different antigen specificities.

A

ANAs

41
Q

The nuclear antigens they are directed against

A
  • double stranded DNA
  • single stranded DNA
  • histones
  • nucleosomes
  • centromere proteins
  • extractable nuclear antigens
42
Q

These antibodies are the most specific for SLE because they are mainly seen in patients with lupus and their levels correlate with disease activity.

A

Double stranded DNA (DsDNA)

43
Q

Antibodies to dsDNA typically produces what type of
staining pattern on indirect immunofluorescence (IIF)?

A

peripheral or homogenous

44
Q

Methods in detecting ANAs have been developed.
⤳ These include the ff:

A
  • Indirect Immunoflurescence test (IIF)
  • Immunoperoxidase staining
  • Enzyme linked immunosorbent
    assay (ELISA)
  • Microsphere multiplex immunoassays (MIA)
  • Radioimmunoassay(RIA)
  • Immunodiffusion
  • Immunoblotting (Western blot)
  • Dot blot
  • Immunoelectrophoresis
  • Microarray.
45
Q

OTHER IMPORTANT INFORMATION
● Note the proper procedure and usage of the __

A

reagents, storage temperature and the expiry date of the kit.

46
Q

Pattern in fana

A

Peripheral (rim)
Homogeneous (diffused)
Speckled
Centromere
Nucleolar

47
Q

Antibodies associated with peripheral pattern

A

Anti-DNA (not seen in HEp-2)

48
Q

Disease associated with peripheral pattern

A

SLE

49
Q

Antibodies associated with homogeneous pattern

A

Anti-DNA
Anti-histone
Anti-DNP (Dinitrophenyl)(nucleosomes)

50
Q

Disease associated with homogeneous pattern

A

Rheumatoid Arthritis
Systemic Lupus Erythymatosus
Miscellaneous Disorders (anti-ssDNA)

51
Q

Antibodies associated with speckled pattern

A

Anti-sm and RNP
Anti- Ro & La
Anti-Jo-1 and Mi-2
Anti- Scl-70

52
Q

Diseases associated with speckled pattern

A

SLE and Systemic Sclerosis (SS)
Polymyositis (PS)/ Dermatomyositis (DM)
PSS (Progressive Systemic Scleroderma)

53
Q

Antibody associated with centromere pattern

A

Anti-centromere

54
Q

Disease associated with centromere pattern

A

Progressive systemic scleroderma
CREST

  • Calcinosis
  • Raynaud’s phenomenon
  • Esophageal dysfunction
  • Sclerodactyly
  • Telengiectaisas
55
Q

Antibody in nucleolar pattern

A

Anti- nucleolar

56
Q

Diseases associated in nucleolar pattern

A

SLE & PSS (Progressive Systemic Sclerosis)

Imse Lab (2 decks)

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