Autoimmunity: ANCA patterns and diseases

Question 1

What disease is associated with the C-ANCA pattern

Answer 1

Wegners granulomatosis

Question 2

What is the target in C-ANCA pattern

Answer 2

Autoantibody targeting Proteinase 3 in the cytoplasm

Question 3

What diseases are associated with P-ANCA pattern

Answer 3

1. Microscopic polyangiitis
2. Churg- Strauss
3. Good pasture syndrome
4. Drug induced ANCA assosicated vasculitis
5. Rapidly progressive glomerulonephritis
6. Inflammatory bowel disease (IBD)
7. PBC
8. Autoimmune hepatitis (type 1)

Question 4

What is the target in P-ANCA

Answer 4

Autoantibodies against myeloperioxidase located in the nucleus

Question 5

Purpose of ANCA IFA

Answer 5

Assist in the diagnosis of suspected ANCA associated vasculitis

Question 6

Principle of method

Answer 6

Diluted patient serum is added to ethnaol fixed human neutrophils. This will form ANCA-neutrophil complexes. After washing off excess or unbound proteins, add anti human IgG FITC (Fluroescein) conjugate which will bind to the Ab-Ag complexes if present. After further incubation and washing complexes are seen under fluorescent microscopy and C-ANCA or P-ANCA can be seen

Question 7

Type of test

Answer 7

Indirect immunofluroscence

Question 8

What is in the pos control for C-anca and P-anca

Answer 8

P-ANCA + control = MPO

C-ANCA = PR3

Question 9

What is the dilution? and Why

Answer 9

1:20
Avoid prozone effect

Question 10

Clinical Significance of ANCA’s

Answer 10

C-ANCA assosicated with Wegners granulomatosis

P-ANCA assosicated with microscopic polyangiitis, idiopathic necrotising and crescentric glomerulonephritis, churg strauss syndrome, good pasture syndrome. AND non vasculitides: IBD, PBC and SLE

ANCA represent a group of autoantibodies directed against cytoplasmic components of the neutrophilic granulocytes.

P-ANCA target antigen = MPO

C-ANCA target antigen = PR3 located in the primary granules

Question 11

Interferences causing false P-ANCA pattern

Answer 11

lactoferrin and elastase and lysozymes

Question 12

Pathophysiology of vasculitis

Answer 12

PR3 and MPO located in the azunophilic granules in the cytoplasm of neutrophils so cant be accessed by Ab.

IL-1 primes the neutrophil and causes the Pr3 and MPO to travel to the surface of the cell membrane

PR3 and MPO can now interact with ANCA in serum

Causes degranulation of Neutrophil leading to ROS formation and thus tissue damage adn THUS vasculitis

Question 13

Why is ethanol used?

Answer 13

The P-ANCA pattern is
an artifact of ethanol fixation, giving rise
to the redistribution of positively charged
antigens to the negatively charged cell
nucleus. If indirect immunofluorescence
is performed using neutrophils fixed in
formalin instead of ethanol, MPO-ANCA
produce a C-ANCA pattern.