Liver antibodies

Liver antibodies mentioned below (except for SLA/LP) are usually screened by indirect immunofluorescence using rodent tissue liver-kidney-stomach.

There are nine types of anti-mitochondrial antibodies which have been identified (namely M1 to M9).  The most commonly detected antibodies are M2 type and highly specific for Primary Biliary Cirrhosis. These can be detected on rodent tissue (liver-kidney-stomach) and/or HEp-2.

Anti-mitochondrial antibodies

Table
Liver


Granular cytoplasmic staining of the Kupffer cells and hepatocytes is characteristic of M2 antibodies.
Kidney

Granular cytoplasmic staining also featured in the renal tubules. Strongest staining is seen in distal which is mitochondria-rich (distal tubules >>proximal tubules).
Stomach

Cytoplasm of gastric cells also appears granular (parietal cells >> chief cells).  
HEp-2 cells

Serum (M2 antibodies) from patients with primary biliary cirrhosis binds to HEp-2 cytoplasm. A fine granular filamentous fluorescence staining is seen.

Antigens: Several anti-mitochondrial antigens exist and the most commonly detected one being M2 which comprises of:

  • 74 kDa E2 subunit of pyruvate dehydrogenase (PDH) complex
  • 55 kDa Protein X of PDH
  • 51 kDa E2 subunit of the branched-chain α-ketoacid/α-ketoglutarate PDH complex
  • 45-50 kDa E1 α-subunit of PDH complex
  • 36 kDa β- subunit of PDH complex

M2 disease association: High titres occur in 95% of patients with Primary Biliary Cirrhosis.

Liver/Kidney Microsomal (LKM) antibodies

LKM-1 antibodies are specific markers of autoimmune hepatitis (type 2).

LKM table
Liver


LKM showing strong homogenous staining of the hepatocytes. Nuclei are not spared. 
Kidney tubules



Renal proximal tubules (large) are preferentially stained but not the distal tubules.
Stomach



There is no staining seen in stomach.


Antigen: Cytochrome p450
Clinical: Type 2 autoimmune hepatitis (80% prevalence)

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Smooth muscle antibodies

ASM
Liver

Antibody reacts with muscle fibres in the walls of the arterial blood vessels.
Kidney

High titre anti-smooth muscle antibody may cross reactivity with tubulin in the glomeruli.
Stomach 

Staining of muscularis propria, intergastric gland area and muscle layers of arteries in the stomach.  
HEp-2 cells   

Cytoplasmic actin staining HEp-2 cells


Antigens: Three major groups of microfilaments are recognised by anti-smooth muscle antibody:

  • Actin (6nm microfilaments)
  • Vimentin and desmin (10nm microfilaments)
  • Tubulin (25nm microfilaments)

Disease association: Autoimmune chronic active hepatitis

Liver cytosolic antigen type 1

Antigen: Identified as the liver specific-enzyme, formiminotransferase cyclodeaminase (62 kDa), which is expression in the cytoplasm but less so around the the central lobular area (see image). LC-1 can coexist with LKM-1 and often masks it. In such cases, it can be resolved with an alternative method such as a liver blot with both antigens. 

Disease Autoimmune hepatitis type 2 (30-50%) in children and young patients.

SLA/LP antibodies

Antigen A 50 kDa UGA-suppressor serine tRNA associated protein. This image was generated by SLA/LP a positive serum obtained from a patient with liver disorder. This may not be representive of SLA/LP antibody but we have seen this associated with SLA/LP serum few times.

The method of choice would be to use a recombinant line blot as shown here.

Disease Severe form of autoimmune hepatitis (type 3?)

Differences between mitochondrial and LKM-1 antibodies

Distinguishing between anti-mitochondrial and anti-LKM-1 antibodies

LKM
 
Antibody
Kidney Stomach Liver
Proximal Distal Parietal cells Chief cells Hepatocytes
M2 ++ +++ ++ -- / + ++
LKM-1 ++ -- -- -- +++
GPC -- -- +++  -- --


From the above table, it is clear that stomach and kidney is required to differentiate between LKM and anti-mitochondrial antibodies.

LKM and gastric parietal cell antibodies

The presence of both LKM and GPC antibodies together can cause a great deal of confusion. Their combined distribution is similar to that of anti-mitochondrial antibodies except for the absence of staining in the distal renal tubules and the presence of smooth homogenous fluorescence in the liver indicating the presence of LKM. This anomaly would have to be investigated further with an alternative method. The example below demonstrates this point clearly.

Liver & kidney
Strong cytoplasmic staining of hepatocytes (negative nuclei) is seen with LKM but in the kidney, only the proximal renal tubules are positive.

 

Kidney
Another image showing positive proximal renal tubules with negative glomerulus.

 

 

Liver & stomach
LKM & GPC staining of gastric parietal cell and liver is common for anti-mitochondrial antibody and often causes confusion.

 

 

Liver/kidney/stomach
Image: lack of staining in the distal renal tubules, which contrasts with mitochondrial antibody. 

This is one of the features which distinguishes between LKM and mitochondrial antibody. Note the additional presence of gastric parietal cell antibody.