Autoimmunity: ANA patterns and diseases

Question 1

What does ANA and Hep2000 stand for

Answer 1

antinuclear antibodies
human epithelioid cells

Question 2

Purpose

Answer 2

Aid in the diagnosis of Rheumatic diseases and in the managment and treatment of these patients

Question 3

Principle

Answer 3

Hep 2000 cells are genetically engineered (human epitheliod cells) with cDNA to express SSA/Ro antigen

Diluted patient serum is incubated with hep2000 substrate allowing specific binding of ab in serum to nuclear material. Incubate to allow any ANA to bind to nuclear antigens forming a stable complex bound to the substrate. Wash off unbound/excess non-specific antibodies. Add fluroescien labelled (FITC) anti human IgG conjugate which will bind to the Ag-Ab complexes. Incubate and wash off excess unbound fluroscein conjugare and observe for patterns and fluroscence level using a Fluroscent microscope.
If the complex is present, a bright apple green fluroscence will occur a staining pattern that is characteristic of the particular nuclear antigen distribution within the cell.

If the sample is negative, there will be a lack of fluroscence in the cell nucleus and the non-chromosome region of mitoitc cells will show brighter staining.

Question 4

Type of test

Answer 4

Indirect immunofluroscnece

Question 5

4 Basic patterns and other patterns and assosicated mitotic cells

Answer 5

1. Homogenous
   fully green cells, mitotic cells have banana of green (chromosome region stained, NO staining of non-chromosome region
2. Speckled
   grainy speckled with black dots, mitotic cells have no staining of chromosome region and non chromosome region has staining
3. Nucleolar
   very few large green clumps surrounded by black, mitotic cells have no staining of chromosome region and non chromosome region has staining

4.Centromere
lots of defined bright green dots, mitotic cells have lines of fluro in chromosome region

Nucleolar and speckled = SSA (base fluro reading off speckled cell fluro level)

Question 6

SLE end point relevance

Answer 6

1:80 or less: rules out active SLE

1:160: Weakly pos often in RA and other connective tissue conditions eg scleroderma, Sjogrens, CAH, TB, maglininacy

1:320 or more: strongly suggests SLE

Question 7

Clinical significance associated to the 4 basic patterns and SSA

Answer 7

Homogenous:
Acute SLE (high titre)
Low titre: SLE and other connective tissue diseases

Speckled:
High titres: SLE, scleroderma, Sjogrens
Lower titres: RA or other connective tissue disease
benign SLE

Nucleloar: High titres: Scleroderma and Sjogrens, SLE

Centromere: CREST, progressive systemic sclerosis

SSA: Primary sjogrens and soemtimes in SLE