Quiz 2 - Autoimmunity & Serological Techniques Flashcards by Fernando Morales

When autoimmunity is suspected, we perform this test…

Anti-nuclear antibodies (ANA)

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Are men or women more predisposed to autoimmune disorders?

Women

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When antigen-antibody complexes deposit in tissue, they attract…

Complement

rupture/lyse cells
autoimmune diseases cause tissue injuries

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3 steps of process of disease

Autoantibodies complex with patient’s own DNA
Immune complexes deposited in vascular systems - organs & tissues
Immune-mediated tissue injury due to complement

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ANA vs Crithidia luciliae - which is the screening test and which is the confirmatory test?

ANA = screening

Crithidia luciliae = confirmatory

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ANA test uses these cells as the substrate

HEP-2

epithelial cell line
ANA reacts with nuclear DNA/RNA

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Why do different ANA patterns appear?

Due to different stages of nuclear DNA mitosis

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How is the ANA test performed?

Patient sera placed on slide with HEP-2 cells and labeled with conjugate antibody with fluorescence tag

ie FITC
“sandwich” assay

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The ANA test is an example of a “sandwich” assay. What is being sandwiched?

Patient’s antinuclear antibodies

-between HEP-2 DNA and conjugate antibody with fluorescence tag

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An ANA test result showing a homogeneous pattern indicates this autoimmune disease

SLE = Systemic lupus erythematosus

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An ANA test result showing a centromere pattern indicates this autoimmune disease

CREST syndrome

-form of scleroderma

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An ANA test result showing a speckled pattern indicates this autoimmune disease

Sjogren syndrome

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An ANA test result showing a nucleolar pattern indicates this autoimmune disease

Scleroderma

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SLE - shows this ANA test pattern. What do the autoantibodies target (3)?

Homogeneous

anti-dsDNA
anti-ssDNA
Histones

-includes anti-Sm abs

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Sjogren - shows this ANA test pattern. What do the autoantibodies target?

Speckled

anti-RNP (ribonuclear protein)

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Scleroderma - shows this ANA test pattern. What do the autoantibodies target?

Nucleolar

anti-nucleolar

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CREST - shows this ANA test pattern. What do the autoantibodies target?

Centromere

anti-centromere

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After performing an ANA test for SLE, we confirm our results with this test. What does the confirmatory test target in the organism?

Crithidia luciliae

Kinetoplast

mitochondria rich in dsDNA
highly sensitive to HEP-2 substrate

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The hereditary component sometimes involved in autoimmune disease involves the…

HLA

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SLE - laboratory findings

Anemia - 50% leukopenia

Thrombocytopenia - 25-50%

Elevated erythrocyte sedimentation rate (ESR)

ANA positive - anti-dsDNA and anti-Sm. Targets nucleoprotein antigens

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Overuse of this causes a SLE-like syndrome. The autoantibodies developed target…

Drugs

Histones
-milder SLE

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SLE vs drug-induced SLE - what would the Crithidia luciliae test results be?

SLE = positive

Drug-induced SLE = negative
-absence of anti-dsDNA antibodies

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SLE - clinical manifestations

Arthritis - chronic inflammation - skin, joints, kidneys, lungs, CNS

Butterfly rash across nose/upper cheeks

Raynaud’s syndrome

Renal disorders

Pulmonary manifestations

CNS disorders - depression, psychosis, convulsions

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What is Raynaud’s syndrome. Which autoimmune disorders is it associated with?

Cyanosis on cold temperatures. Stress-induced vasoconstriction on extremities

SLE, CREST (the “R”)

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Sjogren's syndrome - 90% of autoantibodies are called...

Rheumatoid factor | -anti-RNP associated with ANA stain

Sjogren's syndrome - laboratory findings

ANA - anti-La Labial salivary gland biopsy for definite diagnosis

Dry mouth and dry eyes are symptoms associated with...

Sjogren's syndrome | -affects lacrimal, salivary & excretory glands

Scleroderma has 2 forms

Progressive diffuse Systemic = CREST

What are the acronyms of CREST?

``` Calcinosis - bone formation Raynaud - vasoconstriction of hands/feet Esophageal involvement Sclerodactyly - harden finger skin Telangiectasia - spider veins ```

Scleroderma - lab findings

ANA - centromere pattern (CREST) or nucleolar pattern

Insulin-dependent diabetes mellitus - insulin production is deficient due to...

Destruction of cells of pancreas - congential rubella - HLA

This autoimmune disease has symptoms that overlap to SLE. How are lab results different from SLE?

Mixed connective tissue disease Absence of multiple anti-SM and anti-dsDNA

Ankylosing spondylitis - males or females more prone? What is a common symptom?

Males Fibrosis of bone synovial capsule

Rheumatoid arthritis - primarily affects...

Synovial joints | -swelling, morning stiffness, weight loss, fatigue

Rheumatoid arthritis - lab findings

RF -latex agglutination detects IgM RF Circulating immune complexes ANA - 14-28% of patients

What antibodies are expected for a hepatitis infection?

Anti-smooth muscle

What antibodies are expected for biliary cirrhosis (PBC)?

Anti-mitochondrial

What antibodies are expected for Goodpasture's?

Anti-glomerular basement membrane | -kidney's glomerulus damaged

What antibodies are expected for pernicious anemia?

Anti-parietal cells

IgG subclasses - which ones are autoimmune and which ones are infectious antibodies?

IgG 1 & 3 = infectious IgG 2 &4 = autoimmune

Multiple sclerosis and myasthenia gravis are similar in that they both affect the...

Nervous system - MS - brain, spinal cord, myelin shealth - MG - neuromuscular - do an MRI scan

Hashimoto's thyroiditis vs Graves disease - both have what in common? How are they different

Enlarged thyroid Hashimoto's - hypothyroidism -weight gain, bradycardia, slow metabolism Graves - hyperthyroidism -weight loss, tachycardia, fast metabolism

Pernicious anemia - this is prevented from being absorbed... due to the lack of this...

Vitamin B12 Intrinsic factor

Difference between infectious liver disease and non-infectious liver disease?

Non-infectious (autoimmune) = ANA towards smooth muscle proteins Infectious = no ANA -see anti-viral, anti-bacterial antibodies

Autoimmune liver disease - lab findings

anti-liver antibodies: anti-liver soluble protein (anti-LSP) anti-liver membrane antibody (anti-LMA) anti-acidoglycoprotein receptor (anti-ASCP) No anti-dsDNA

The highest peak on a normal SPE graph corresponds to...

Albumin

Multiple myeloma patients show a large spike in the SPE corresponding to...

gamma-globulins

What does IFE stand for? What is the test used to determine?

Immunofixation Electrophoresis Determines what immunoglobulin abnormalities exists

After band separation, IFE uses ___ to probe for immunoglobulins and visualize bands

Anti-antibodies

Complement can be inactivated by ___

Heat | -56C for 30 mins

The complement system is activated by...

Antibodies

Name two assays to detect complement

ELISA CH50

The CH50 assay is used to detect hemolytic activity of... What does the CH50 value mean?

Complement Amount of patient serum required to lyse 50% of a standard concentration of sheep RBC

How is titer calculated from a dilution?

Titer is the reciprocal of the dilution

Strep toxin can cause this substrate, ___, to change from blue to pink. What does strep toxin do? If patient has antibodies against strep toxin, what color will the test be?

Toluidine blue Depolymerize DNA = pink color Blue - toxin neutralized - no color change

How is the dilution calculated from the dilution factor?

Reciprocal | -dilution factor 5 is 1:5 dilution or 1/5

Antigen-antibodies can form 3 zones depending on their concentration. What are the 3 zones as it relates to the amounts of antibodies and antigen?

Prozone = too much antibody Zone of equivalence Postzone = too much antigen

Precipitation test uses the concept of... using this media...

Immunodiffusion Agar plate

3 types of agglutination tests

Latex agglutination - mono, RA, rubella Hemagglutination Inhibition reactions

Difference between turbidimetric and nephelometric

Turbidimetric = measure light absorbance by immune complex Nephelometry = measure light scatter by immune complex

Immunodiffusion assays have this weakness. However, they're still useful for detecting ___ infections

Takes a long time to do Fungal

A double immunodiffusion assay is also called... What are the patterns that form?

Ouchterlony Identity Partial Identity Non-identity

Describe what is happening in each Ouchterlony assay reaction

Identity - both antigen/antibody and antigen/unknown have matching epitopes and will diffuse radially towards one another Partial identity - share a few epitopes, but have more not in common. Forms small bifurcation/spur Non-identity - precipitation lines cross, do not share epitopes

Radial Immunodiffusion (RID) - describe assay. What does the result look like?

Antibody infused into agar. Inoculate patient sample into well and let diffuse out to zone of equivalence Zone of equivalence = precipitation ring -ring diameter proportional to concentration

IEP vs IFE - which is more sensitive and produce sharper bands? How are they similar?

IFP Both use electrophoretic immunodiffusion

Terms for when Igs in a patient are lower than normal, higher than normal, and none

Hypogammaglobinemia Hypergammaglobulinemia Agammaglobulinemia

Agglutination tests that use RBCs in the assay are called...

Hemagglutination tests

Agglutination assays can be direct or indirect. What does each one mean?

Direct - antigen-antibody bind directly Indirect - antigens are coated on a carrier molecule -makes reaction more visible, easier to interpret

Flocculation assay is a subset of this type of test... It is rarely used besides detection of this disease...

Agglutination -precipitation of fine particles Syphilis -VDRL and RPR

Describe the complement fixation test. Describe the positive and negative results

Complement is "fixed" with antigen-antibody so it cannot react and lyse RBCs -uses sheep RBCs ``` Positive = antigen-antibody complex present, no lysis Negative = antigen-antibody complex not present, lysis ```

Immunoblot - difference between southern blot and western blot

Southern = DNA Western = protein

Define heterophile antibodies. An example of a heterophile antibody is known as...

Antibodies that react with antigens not responsible for their production = non-specific Forssman antibody - IgM in nature - made against guinea pig kidney but reacts with sheep RBCs

2 different tests to detect heterophile antibodies

Paul Bunnell heterophile antibody test Davidsohn differential test