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.
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 a-ketoacid/a-ketoglutarate PDH complex
- 45-50 kDa E1 a-subunit of PDH complex
- 36 kDa ß- 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 showing strong homogenous staining of the hepatocytes. Nuclei are not spared.
Renal proximal tubules (large) are preferentially stained but not the distal tubules.
There is no staining seen in stomach.
Antigen: Cytochrome p450
Clinical: Type 2 autoimmune hepatitis (80% prevalence)
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Smooth muscle antibodies
Antibody reacts with muscle fibres in the walls of the arterial blood vessels.
High titre anti-smooth muscle antibody may cross reactivity with tubulin in the glomeruli.
Staining of muscularis propria, intergastric gland area and muscle layers of arteries in the stomach.
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.
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
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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.
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.
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.