Immunology Testing Flashcards
What are the four ways an ANA can be helpful?
- help diagnose a patient who is suggestive of an AI disease
- hep exclude AI disease
- subclassify patients with AI
- monitor disease activity
What is the sensitivity of a positive ANA for SLE?
95% - pretty good
less so for other autoimmune diseases - not specific at all
What are some non-AI causes of a +ANA?
Mono, hepC, lymphoproliferative disease, HIV, anti-synthetase syndrome (cytoplasmic staining)
How is an ANA run?
A HEp2 (human epithleial cell tumor line) is used with acetone fixed substrate (more sensitivie)
you get a report pattern and titer (1:80 is considered significant). But be cautious: up to 20% of people will have a normal titer over 1:80 and some may even have a normal titer over 1:320. So take in clinical context.
What pattern of +ANA testing is usually seen in SLE?
homogeneous pattern
Describe this homogeneous pattern.
You get positive staining of the cells while cytoplasm is negative - this is because the stain divides the cell chromatin.
What are the two antibodies that give rise to this homogeneous pattern?
Anti-DNA histone complex
Anti-dsDNA
What are the two strategies used to identify this Anti-dsDNA?
C. lucilliae
ELISA (mostly taking over - more sensitive, but not as specific)
Antibodies to extractible nuclear antibens will result in what +ANA pattern?
speckled
What are some examples of these “extractible nuclear antigens”?
Sm, RNP, Ro/SSa, La/SSb, Scl-70, centromere, PCNA
Anti-smith is very specific for SLE patients, but only ___% of SLE patients will have it.
25
Anti-U1nRNP is suggestive of what diagnosis?
mixed connective tissue disease
What symptoms are associated with antibodies to RO/SSA in SLE?
photosensitivity sicca thrombocytopenia subacute cutaneous LE rash Neonatal lupus (maternal IgG crosses placenta - can lead to complete heart block and pregnancy loss!)
The Nucleolar pattern of +ANA has various specificities and an also be present in in 20% of people with what disease?
scleroderma
The centromere +ANA pattern is from antibodies against what?
proteins present on active centromeres during meiosis and mitosis: CENP-A, CENP-B, CENP-C
What disease is the centromere +ANA pattern associated with?
a CREST variant with calcinosis and telangiectasis but less renal invovlement
What are the two strategies for identification of the specific antibodies involved in a +ANA?
ELISA or multiplex
Describe multiplex testing
uses colored polystyrene beads coated with single purified antignes
uses fluorescein-labeled antihuman IgG that is read in a dual-laser flow cytometer
What is the primary use for ANCA testing?
evlauate patients with suspected vasculitis (usually either granulomatosus with polyangiitis or microscopic polyangiitis)
cANCA is usually due to anti-_____
anti proteinase 3
pANCA is usually due to anti-____
myeloperoxidase
Granulomatosus with polyangiitis is usually associated with which one?
cANCA
So microscopic polyangiitis is associated with…
pANCA
What are the criteria for diagnosis of RA?
Need 5 of the following: morning stiffness pain on motion joint swelling symmetrical joint involvement subQ nodules x-ray changes histologic changes on biopsy \+RF \+Anti-CCP
What is rheumatoid factor?
Usually IgM antibody that reacts with the Fc portion of IgG (all, not just human)
Describe the rheumatoid factor test.
Usually done with ELISA (often with rabbit IgG)
can also do it with latex particles coated with non-human or human IgG, which will agglutinate the RF is present; quantitation done by titering the antibody.
WHat are some other conditions with RF present?
bacterial endocarditis syphilis hep B and C infectious mono myeloma cryoglobulinemias sarcoidosis
What is cyclic citrullinated peptide derived from?
fillagrin, which is associated with keratin filaments
True or false: +anti-CCP is predictive of milder disease.
false - predictive of more severe disease
WHat is the positive predictive value of CCP and RF together?
over 97% - huge
What are the main cytokines released in RA?
IL-6, IL-12, TNF-a, IL-15
What are some clinical manifestations of celiac disease?
chronic diarrhea, weight loss, malabsorption, abdominal distension (typical)
dermatitis herptiformis
osteoporosis (atypical)
short status (atypical)
neuropsychiatric (dep, anx, ataxia, migraines, neuropathy, etc.)
What is the incidence for celiacs?
1:100 (wow)
What do you see on GI biopsy in celiacs?
villous atrophy
Describe the pathophysiology of celiac disease.
gluten-reactive CD4+ T cells in the lamina propria of the upper GI tract react with the alpha-gliaden peptide sequence in gluten
presents peptides by HLA-DQ2 or 8, which triggers production of proinflammatory cytokines
tissue transglutaminase deaminates the peptides, and the deaminated glutamine peptides then bind more avidly to HLA sites, further increasing immunogenicity
enterocytes produce IL-15 which activates NK cells
all leading to villous atrophy, crypt hyperplasia and intraepithelial lymphocytosis
We use IgA anti-endomysial antibodies as a highly specific test for CD. What does it bind?
tissue transglutaminase
Why is tissue transglutaminase a good antigen to test for in CD?
it’s released from intestinal mucosa with injury and also plays a role in the pathogenesis of disease (deaminates glutamine to glutamic acid, which binds more avidly to HLA-DQ2 and 8 as before)
Which immunodeficiency is much more common in CD than in the general population?
IgA deficiency
What test is the most efficient for making a diagnosis of CD?
IgA anti-tissue transglutaminase
What test should you run to make the diagnosis of CD in IgA deficient patients? (that is if they’re otherwise low risk - lots of false positives in moderate to high risk patients)
IgG anti-deaminated gliadin
What are the two common HLA haplotypes in CD?
Hinting at them before: HLA-DQ 2 (95%) and 8 (5%)
absence has high negative predictive value and virtually excludes CD
What is the best antibody to monitor treatment in CD?
anti-tTG
normal levels will be reached after 3 to 12 months of gluten free diet
What antibodies are associated with Type I Autoimmune hepatitis?
ANA anti-SM anti- soluble liver antigens anti-saccharomyces cerevisiae anti asialoglycoprotein receptor pANCA
What antibodies are associated with Type 2 autoimmune hepatitis?
anti liver/kidney microsomes (cyt p450)
anti liver cytosol antigen
Type 1 autoimmune hepatitis is associated with what HLA haplotype?
DR3
Type 2 autoimmune hepatitis is associated with what HLA haplotypes?
DRB1
DQB1
Anti-smooth muscle antibodies come in different flavors specific to specific parts of myocytes. WHat’s the most common one in autoimmune hepatitis?
anti-actin antibodies (this is more sensitive and specific than the general anti-smooth muscle antibodies)
pANCA is associated with type 1 autoimmune hepatitis, but instead of being directed against myeloperoxidase as it is in the vasculitides, what is the target?
myeloid 50kD nuclear envelope protein
note: this atypical pANCA is also found in ulcerative colitis
How does primary biliary cirrhosis present?
usually women ages 30-65 with fatigue, pruritus, hyperpigementation/jaundice, arthropathy common,
sjogrens in 50%
increased alk phos, slight increase in aminotransferases, HLD
What antibody is associated with PBC?
anti-mitochondrial antibodies
also ANA in 30%
What are the four autoantigens that anti-mitochondrial antibodies go after?
E2 subunit of PD
2-oxo-acid dehydrogenase
ketoglutaric acid dehydrogenase
hydrolipoamide dehydrogenase
(all involved in oxidative phosphorylation within the mitochondria)
