IMMUNOLOGY SERVICES
SCOTTISH NATIONAL
BLOOD TRANSFUSION
SERVICES
EDINBURGH
USER MANUAL; V2.2
Revised April, 2008
HTTP://WWW.SCOTBLOOD.CO.UK/DOCS/IMMUNOLOGY.DOC
CONTENTS
GENERAL INFORMATION p3
AUTOANTIBODIES IN ENDOCRINE DISORDERS p4
AUTOANTIBODIES IN LIVER DISEASES p7
AUTOANTIBODIES IN PRIMARY VASCULITIDES & p9
GOODPASTURE‟S SYNDROME
AUTOANTIBODIES IN CONNECTIVE TISSUE DISEASES p10
MISCELLANEOUS AUTOANTIBODIES p16
GUIDELINES FOR REPEAT TESTING OF AUTOANTIBODIES p19
ANA Q&A p20
MEASUREMENT OF COMPLEMENT PROTEINS p21
INVESTIGATIONS OF IMMUNODEFICIENCIES p25
GUIDELINES FOR ALLERGY TESTING p27
SPECIFIC IgG ANTIBODY TEST AGAINST FUNGAL & AVIAN p29
PRECIPITINS
GENERAL INFORMATION
BTS Immunology Laboratory,
Edinburgh & SE Scotland Blood Transfusion Service,
Royal Infirmary of Edinburgh,
51 Little France Crescent,
Edinburgh, EH16 4SA.
Opening Times
Monday – Thursday: 8.30 am to 5.00 pm
Friday: 8.30 am to 4.30 pm.
There is no weekend working or out of hours service.
Blood Samples
1 Specimens should be labelled with the patient's full name, date of birth, ward, outpatient department or general
practice, and date of collection. The patient's CHI number must be given where available and Addressograph
labels may be used.
2 Specimens should be accompanied by the appropriate Request Form completed, including all patient details as
stated in para 1 above, relevant clinical information and previous report numbers (if available), signed by a member
of the medical staff. It is essential to indicate where the Report of Test Results should be sent to.
3 Specimens that are incorrectly labelled or documented may not be accepted.
This laboratory has the responsibility for the investigation of patients suspected of suffering from disease
consequent on abnormal function of the immune system, in particular, autoimmunity, immune deficiency and
allergy.
Tests for Autoantibodies, Complement, total IgE & RAST (specific IgE tests)
5-10 ml of clotted blood should be sent in an anticoagulant free tube.
For children, a 1-2ml clotted sample may be sent.
Transport
All blood samples should be sent to the above address either by post, following current guidelines, or by using the
LUHT delivery service.
Reporting of Results
All test requests are handled as expeditiously as possible but some specialised tests are redirected to other centres.
Results can be obtained by telephoning the laboratory when needed urgently.
The Consultant Clinical Immunologist is available by telephone for discussion of the clinical implications of test
results
Staff and Telephone Numbers
Consultant Clinical Immunologist…………….……………………………………………. Dr PL Yap
Head of Immunology Laboratory…..………………………………………………………. Mrs J Sawers
Deputies …………………………………….……………………………………………… Mrs G Steven
……………………………………………………………………………………. Ms G McNab
Prinicpal Clinical Scientist…………………………………………………………………. Dr M Kadlubowski
Immunology Laboratory, Reception & Results…………………………………………….. 0131 242
7525/7529
…………………………………………………………………………………….. (Internal 27521/27529)
Dr PL Yap………………………………………………………………………………….. 0131 242 7526
…………………………………………………………………………………….. (Internal 27526)
Dr Yap‟s Secretary…………………………………………………………………………..0131 242 7521
…………………………………………………………………………………….. (Internal 27521)
Abbreviations used:
IFF – Indirect Immunofluorescence
ELISA – Enzyme Linked Immunosorbent Assay
AUTOANTIBODIES IN ENDOCRINE DISORDERS
Anti-Thyroid Peroxidase Antibodies (anti-TPO)
Introduction
Thyroglobulin (MW 660 kD) is the major protein of the colloid of the thyroid follicular cells. Thyroid peroxidase
(TPO), the thyroid microsomal antigen, is a 110kD peroxidase enzyme found on intracellular membranes of the
epithelial cells of the follicles and its function is to incorporate iodine onto the benzene ring of thyroglobulin
tyrosine residues. In response to thyroid stimulating hormone, iodinated tyrosines are cleaved by lysosomal
proteases to eventually yield thyroxine (T4) and triiodothyronine (T3).
Autoantibodies to TPO are predominantly directed against 2 immunodominant regions which reside in a region
with structural homology to myeloperoxidase. They are frequently found in very high levels in sera of patients with
autoimmune thyroid disease Graves‟ disease, Ha\shimoto‟s thyroiditis and primary atrophic hypothyroidism).
They are usually IgG and rarely respond to treatment. Autoimmune thyroiditis may be associated with other
autoimmune diseases especially pernicious anaemia or myasthenia gravis.
Use of Tests
Diagnosis of autoimmune thyroid disease.
Method Used
ELISA for IgG anti-TPO.
Reporting of Results
Results are expressed as IU/ml where >82 IU/ml is positive. A borderline result is in the range of 42-81.9 IU/ml.
High levels (>5000 IU/ml) can be detected in patients with autoimmune thyroiditis. The turnaround time is 5
working days.
Interpretation of Results
Anti-TPO antibodies are detected in raised levels (>82u/ml) in:
85-100% of patients with overt autoimmune thyroiditis (Hashimotos thyroiditis)
70-90% of patients with Graves disease
60% of patients with primary hypothyroidism
30-60% of patents with pernicious anaemia but no overt thyroid disease
Anti-thyroid antibodies are also frequently found in patients with other autoimmune diseases eg
Pernicious anaemia 60%
Myasthenia gravis 50%
Lambert-Eaton syndrome 30%
They are detected in low levels in 2-8% of normal individuals particularly the elderly and more often in women
than men.
Anti-thyroglobulin autoantibodies (anti-Tg) are no longer assayed. However, this can be arranged by special
request in patients with suspected thyroid cancer.
It is very unusual to detect anti-Tg antibodies in the absence of anti-TPO antibodies but quite common to find
anti-TPO antibodies in the absence of anti-Tg antibodies, especially in patients with small goitres.
Anti-Tg requests are sent to an external laboratory for testing* (see p 18 )
Anti-Gastric Parietal Cell Antibodies (anti-GPC or PCA))
Introduction
The parietal cells of the gastric mucosa secrete a protein, intrinsic factor, which is essential for the absorption of
vitamin B12, a vitamin needed for normal erythropoiesis. The target antigen for GPC is the transmembrane
ATP-dependent proton pump. The latter consists of an alpha catalytic subunit MW approx. 90 kDa) and a beta core
subunit (MW approx. 35 kDa). The resulting destruction of the parietal cells leads to inability to absorb the
vitamin B12 and hence to anaemia. Antibodies against gastric parietal cell antigens are diagnostic of chronic
autoimmune gastritis whether or not the result is pernicious anaemia. Very severe gastric atrophy is necessary
before anaemia is detected. . Antibodies to intrinsic factor can also be detected in the same diseases.
Anti-gastric parietal cell antibodies (in the presence or absence of anaemia) are frequently associated with other
autoimmune diseases such as autoimmune thyroiditis, insulin dependent diabetes or Addison‟s disease.
Use of Test
Diagnosis of pernicious anaemia.
Method Used
Indirect immunofluorescence (IIF) on rat gastric mucosa.
Reporting of Results
Positive results are expressed from +/- (a borderline result) to +++ (strongly positive). The turnaround time is 7
working days.
Interpretation of Results
Anti-gastric parietal cell antibodies are found in >90% patients with pernicious anaemia. They are not associated
with duodenal ulcer or gastritis which is not autoimmune. Anti-gastric antibodies are also found in:
30% patients with Addisons disease
30% patients with autoimmune thyroiditis
30% patients with insulin dependent diabetes
20% patients with primary ovarian failure
Around 5% of normals have low levels of these autoantibodies rising to 10% in older women. They are more
common in relatives of patients with autoimmune thyroid disease or gastritis. The antibodies do appear many
years before the onset of pernicious anaemia and therefore patients with high elevated levels should be followed
up.
Anti-Intrinsic Factor Antibodies (IFA)
Two types of IFA are recognised. Type I blocks the binding of vitamin B12 to intrinsic factor and Type II prevents
attachment of intrinsic factor to the ileal mucosa. Both types are measured in the one method in the laboratory.
These antibodies are specific for patients with pernicious anaemia but are only found in 50-60% of patients with
pernicious anaemia.
These tests are performed infrequently and only on patients with a low serum B12 level and high red cell mean
corpuscular volume.
Method Used
ELISA for IgG IFA.
Reporting of Results
Results are reported as „Negative‟ ' Borderline' or 'Positive'. The turnaround time is 10 days.
Anti-Adrenal Cortex Antibodies/Anti-Steroid Cell Antibodies
Introduction
Antibodies to the adrenal cortex usually stain the glomerula, fasciculate and reticulosa cells and are directed against
a number of enzymes involved in steroid metabolism (esp. Steroid 21-Hydroxylase, Steroid 17-alpha Hydroxylase
and a number of Cytochrome P450 subtypes) . Such antibodies have been detected by a variety of procedures, but
the method most commonly used is IIF. The antibodies belong predominantly to the IgG class.
Use of Test
Diagnosis of Addison‟s disease.
Method Used
IIF using adrenal tissue sections, using a goat anti-human IgG.
Reporting of Results
Positive results are expressed from +/- (a borderline result) to +++ (strongly positive). The turnaround time is 10
working days.
Interpretation of Results
Anti-adrenal cortex antibodies are found in:
75% of patients with Addison‟s disease at the time of diagnosis.
Also found occasionally in IDDM and autoimmune polyendocrinopathies...
They are not found in tuberculosis associated adrenal failure or secondary carcinoma. They are very rare in normal
individuals (0.6%), usually in older women.
Anti-Pancreatic Islet Cell Antibodies (ICA)
Introduction
Islet Cell antibodies (ICA) occur in 65-85% of patients with Type I (insulin-dependent, juvenile) diabetes mellitus
early in the course of the disease. The standard method for the detection of these antibodies is IIF. Islet cell
antibodies, when detected by IIF, stain the cytoplasm of islet cells of the pancreas from humans, monkeys and rats.
A large number of target antigens to ICA have been identified and these include glutamic acid decarboxylase
(GAD), protein tyrosine phosphatase (ICA512), phogrin and glycolipids. 60 to 80% of individuals with new onset
insulin dependent diabetes mellitus have anti-GAD antibodies.
Use of Test
Diagnosis of Type I, insulin dependent/juvenile, diabetes.
Method Used
IIF using monkey pancreas sections.
Reporting of Results
Positive results are expressed from +/- (a borderline result) to +++ (strongly positive). The turnaround time is 10
working days.
Interpretation of Results
Anti-islet cell antibodies are present in 65-85% of patients with Type I diabetes particularly around the time of
diagnosis. They tend to disappear later unless there is multi-endocrine disorder. They are also found less
commonly in non-diabetic relatives.
AUTOANTIBODIES IN LIVER DISEASES
Anti-Smooth Muscle Antibodies (SMA)
Introduction
Sera from patients with Type 1 chronic active hepatitis contain antibodies to smooth muscle antigens that are
detectable by IIF and stain the smooth muscle of several organs. The antibodies belong mainly to the IgG class,
but they can also be found in the IgM class. The antigen recognised by anti-smooth muscle antibodies in sera of
patients with Type 1 chronic active hepatitis is often reported to be actin. However, this is a very complex and
incompletely characterised group of autoantibodies which in liver disease and in other diseases can recognise
desmin, vimentin, tropomyosin, cytokeratins or other cytoskeletal proteins.
Use of Test
Anti smooth muscle antibodies can be diagnostic of Type 1 chronic active hepatitis and are useful in differential
diagnosis of liver diseases.
Method Used
IIF using rat liver, kidney and stomach tissues.
Reporting of Results
Positive results are expressed from 1:10 (a borderline result) to >1:640 (strongly positive). The turnaround time is
7 working days.
Smooth muscle antibodies are detected in:
40-70% of patients with Type 1 chronic active hepatitis
50% of patients with primary biliary cirrhosis
28% of patients with cryptogenic cirrhosis
Type 1 chronic active hepatitis is often associated with hypergammaglobulinaemia and with anti-nuclear
antibodies.
Anti-smooth muscle antibodies are also found in patients with acute viral hepatitis, infectious mononucleosis,
asthma, yellow fever and malignant tumours (carcinomas of the ovary, malignant melanoma). They have been
found in less than 2% of the normal population. Antibody titres are in the range of 80 to 320 in Type 1 chronic
active hepatitis and are much lower, rarely over 80, in the other conditions listed above. They can be found in
patients with rheumatoid arthritis, multiple sclerosis but probably not SLE.
Anti-Mitochondrial Antibodies (AMA)
Introduction
Anti-mitochondrial antibodies are frequently associated with autoimmune liver disease. There are a number of
antibody specificities which have been reported. These are termed M1-M9. M2 (the subtype reported here) are
the most common ones observed and these are highly specific for Primary Biliary Cirrhosis. The M2
autoantibodies recognise mainly four closely related enzymes in the 2-oxoacid dehydrogenase complex; pyruvate
dehydrogenase, branched chain 2-oxoacid dehydrogenase, 2-oxoglutarate dehydrogenase and an unknown protein.
Most antibodies recognise the 74kDa acyltransferase E2 component of the enzymres and the related 45kDa protein.
These autoantibodies are most frequently associated with primary biliay cirrhosis. Antibodies of other specificity
are associated with other liver diseases. The other 8 specificities recognise a disparate number of antigens and are
observed in a variety of disorders including syphilis (M1), coonective tissue diseases (M5) and cardiomyopathies
(M7).
Use of Test
Diagnosis of primary biliary cirrhosis; differential diagnosis of liver disease.
Method Used
IIF using sections of rat liver, kidney and stomach.
Reporting of Results
Positive results are expressed from 1:10 (a borderline result) to >1:640 (strongly positive). The turnaround time is
7 working days.
Interpretation of Results
Anti-mitochondrial antibodies of the M2 type are present in at least 90% of patients with PBC but can also be
o
present in other autoimmune disease such as 2 Sjögrens syndrome, autoimmune thyroid disease, SLE or
polymyositis. Can be associated with anti-smooth muscle antibodies in PBC/CAH overlap.
Anti-Liver/Kidney Microsomal Antibody (anti-LKM1)
Antibodies recognising this antigen are found in a rare subset of patients with Type 2 chronic active hepatitis
occurring usually in young patients. The antibodies react with a 50-57kDa protein (Cytochrome P450 2D6) found
on cells in the third portion of the proximal renal tubules and on hepatocytes. They may also react with cells in
bronchial, oesophageal and duodenal epithelium. These autoantibodies are not associated with ANA or SMA
which tend to be associated with the more common Type 1 chronic active hepatitis.
There are also two much rarer LKM subtypes. LKM2 targets other cytochrome P450 subtypes and is associated
with drug-induced hepatitis and LKM3 targets UDP-glucuronyltransferase and is associated with Hepatitis D.
The much rarer LKM2 autoantibody is associated with drug-induced liver disease.
Use of Test
Diagnosis of Type 2 chronic active hepatitis in the young.
Method Used
IIF using sections of rat kidney, liver and stomach sections
Reporting of Results
Results are expressed as either „Positive' or „Negative' The turnaround time is 7 working days.
Interpretation of Results
A positive result is diagnostic of this subset of patients with Type 2 chronic active hepatitis.
L/K microsomal antibodies are also found in patients with chronic active hepatitis due to hepatitis C infection -
usually in middle aged males of Mediterranean extraction.
AUTOANTIBODIES IN PRIMARY VASCULITIDES & GOODPASTURE’S
SYNDROME
Anti-Neutrophil Cytoplasmic Antibodies (ANCA)
Introduction
Autoantibodies recognising antigens in the cytoplasmic granules of neutrophils (ANCA) were first detected in a
number of patients with Wegener‟s granulomatosis. There are two clinically relevant staining patterns found in
ethanol-fixed granulocytes known as c-ANCA and p-ANCA. c-ANCA is a granular cytoplasmic staining pattern
which in approx. 50% of cases is due to recognition of Proteinase 3 (PR3), a primary granule 29kDa protease.
p-ANCA is a perinuclear staining pattern due to recognition of Myeloperoxidase (MPO), Cathepsin G, Elastase and
other antigens which relocate to the perinuclear region during ethanol fixation of the cells. Other
immunofluoresence staining patterns termed atypical-, x- and a-ANCA have been reported and are found in a wide
range of diseases including vasculitides, connective tissue diseases, inflammatory bowel diseases and infections.
They are the result of antibodies to at least 20 neutrophil proteins. Of these only autoantibodies to proteinase 3
(PR3-ANCA) and myeloperoxidase (MPO-ANCA) are considered to be of diagnostic and prognostic usefulness.
The latter are measured by ELISA using purified human antigens.
Use of Test
The diagnosis of primary vasculitides (mainly small vessel) such as Wegener‟s granulomatosis and microscopic
polyangiitis and the monitoring of treatment.
Method Used
ANCAs are measured by IIF and/or an ELISA screen (which measures IgG antibodies to both PR3 and MPO).
Both of these methods are used to screen patient samples. All positive samples are then tested for ANA by IIF (see
below) and PR3-ANCA and MPO-ANCA by ELISA.
Reporting of Results
IIF Results are expressed as pANCA, cANCA, atypical pANCA, atypical cANCA, atypical ANCA or negative.
ELISA screen results are expressed as negative, equivocal or positive. All equivocal and positive results are further
assayed for IgG antibodies to PR3-ANCA and MPO-ANCA. The turnaoround time is 10 working days.
Borderline 5.0 to 9.9 AU/ml
Positive >10 AU/ml
Interpretation of Results
It is important to be aware that in an unselected population barely half of c-ANCAs are caused by autoantibodies to
PR3 and that only ~25% of p-ANCAs are caused by autoantibodies to MPO. It should therefore never be assumed
that c-ANCA and p-ANCA results are synonymous with PR3-ANCA and MPO-ANCA respectively. The majority
of patients with Wegener‟s granulomatosis (WG) and microscopic polyangiitis (MPA), however, will have
antibodies to PR3 or MPO. PR3-ANCA are much more common in WG whereas MPO-ANCA are more common
in MPA.
ANCAs are also found in patients with segmental necrotising glomerulonephritis and in patients with crescentic
glomerulonephritis. IgA anti-neutrophil antibodies have been detected in Henoch-Schönlein purpura associated
with IgA rheumatoid factor.
Atypical ANCAs not due to PR3 or MPO antibodies are often found in patients with inflammatory bowel disease.
The antibodies occur in some patients with ulcerative colitis and less commonly in patients with Crohn's disease
with colon involvement and also in patients with sclerosing cholangitis associated with Crohn's disease.
Granulocyte specific anti-nuclear antibodies are found in rheumatoid arthritis and must not be confused with
ANCAs.
Anti-Glomerular Basement Membrane Antibodies (anti-GBM)
Introduction
Antibodies recognising glomerular (and alveolar) basement membranes are characteristic of Goodpasture‟s
syndrome. The antigen is most frequently the non-collagenous domain (NC1) of the alpha 3 chain of collagen IV.
These anti-glomerular basement (GBM) antibodies are pathogenic and are usually IgG. There appears to be some
subclass restriction with IgG1 and IgG4 predominating.
Use of Test
Diagnosis of Goodpasture‟s Syndrome.
Method Used
ELISA for IgG anti-GBM.
Reporting of Results
Results are expressed as AU/ml where > 20 is positive. The turnaround time is 10 working days.
Interpretation of Results
Anti-GBM antibodies are found in the sera of the majority (>90%) of patients with Goodpasture‟s syndrome. The
diagnosis must be confirmed by renal biopsy. They are also occasionally found in ANCA-associated disease (see
above), usually with MPO-ANCA but very rarely with PR3-ANCA.. Less than 2% of patients with
glomerulonephritis have these autoantibodies.
ANCA/GBM Dot-Blot
An emergency qualitative immunoblot assay for antibodies to PR3, MPO and GBM is available by request only
with the approval of the consultant immunologist. The blood sample must reach the immunology laboratory by
1.00 pm at the latest . The result will be telephoned to the requesting physician between 4.00 and 4.30 pm on the
same day and will be reported as positive or negative for each of the 3 specificities. Any positive result will be
confirmed by the appropriate ELISA as above.
AUTOANTIBODIES IN CONNECTIVE TISSUE DISEASES
Anti-Nuclear Antibodies (ANA)
Introduction
The detection of circulating antibodies to nuclear antigens is an important tool in the investigation of systemic
connective tissue diseases. Many techniques have been developed to detect antinuclear antibodies (ANA), but the
fluorescent-ANA (FANA) test continues to be the most widely used and accepted. We and many other
laboratories use this test to screen sera before other techniques are used to define antibody specificity. Compared
with these more elaborate techniques, the FANA test has the advantages of broad specificity, sensitivity, economy,
reproducibility and relative ease of performance. A wide range of ANAs are detected by this technique because
most nuclear antigens are represented in carefully prepared tissue substrates. The ANA of patients with systemic
connective tissue diseases are not restricted by tissue specificity and will therefore bind to nuclear components from
various species. Exceptions to this rule include sera that react specifically with human leukocyte nuclei and to
Sjögren's syndrome antigen A (SS-A/Ro).
Use of Test
The major reason for ordering the FANA test is to confirm the clinical diagnosis of a systemic connective tissue
disease such as systemic lupus erythematosus (SLE). A negative FANA test does not completely rule out the
diagnosis of SLE, but alternative diagnoses should be considered. Anti-nuclear antibodies, particularly at low
titres are associated with a number of autoimmune diseases. In addition, patients receiving a drug such as
procainamide, phenytoin or hydralazine, should be tested if symptoms occur suggesting a diagnosis of drug-induced
lupus erythematosus.
Method Used
Indirect immunofluorescence (IIF) is performed on fixed Hep2000 cells. The titre of antibody and the pattern of
staining is reported. This cell line has large nucleii which allows better definition of the staining pattern and
preparations contain approx. 10% of cells engineered to contain elevated levels of Ro60. There are also significant
numbers of dividing cells allowing detection of autoantibodies to cell cycle specific antigens such as centromere
proteins.
Reporting of Results
Results are reported as 'Negative' or 'Positive' with the antibody titre on HEp2000 cells reported as 1:40
(borderline) to >1:640 (strongly positive). The turnaround time is 5 working days.
The intensity of staining depends on the antigen which is being detected. This also has a marked effect on the
staining pattern. Although a large number of staining patterns have been described, we classify the staining
patterns into various groups. The major staining patterns are:
Homogeneous Homogeneous staining of interphase nuclei plus positive staining in the chromosomal region of
mitotic cells
Speckled Large, coarse or fine speckled staining of the interphase nuclei with negative staining in the
chromosomal region of mitotic cells
Homogeneous/Speckled Very similar staining pattern to speckled but with positive staining of the chromosomal
region of mitotic cells.
Atypical Speckled This staining pattern is specific to HEp2000 and results from the fact that approx. 10% of the
cells present are engineered to contain significant amounts of Ro. It is only these 10% of cells showing nucleolar
and a very fine speckled pattern in the rest of the nucleus.
Nucleolar Staining of nucleolus only. The appearance can vary depending on which of several nucleolar
antigens are involved
Homogeneous/Nucleolar Staining of the nucleoli only of interphase nuclei together with positive staining in the
chromosomal region of mitotic cells.
Centromere Approx 40 discrete dots caused by staining of the centromere regions of the chromosome which
interact with the mitotic spindles. The dots are found in the condensed nuclear chromatin during mitosis. The
centromere antigen is located at the point of attachment of the chromosome to the mitotic spindle. The major
antigen recognised is an 80kDa DNA binding protein termed CENP-B.
Nuclear Membrane Similar to homogeneous but with more intense staining of the nuclear membrane (also called
rim staining)
Many minor staining patterns are observed and these include Mitotic Spindle, PCNA, Centriole, Nuclear Dots and
a large number of mixed patterns (ie. 2 of the above in the same patient sample).
Interpretation of Results
All positive ANAs (>10) are titred out (40, 160, 640)
A Positive ANA may trigger the measurement of anti-dsDNA if requested (dependent on pattern and titre).
A homogenous/nucleolar pattern triggers an ENA screen and a positive ATS pattern will trigger an ENA profile.
The pattern of IIF will often provide a clue to the category of nuclear antigens involved. For example, antibodies
binding nonhistone proteins (Smith [Sm], nuclear ribonuclear protein [nuclear RNP], La antigen [SS-B]) give a
speckled pattern of immune fluorescence, antibodies to DNA and histone give a homogenous or rim pattern, and
antibodies to 4 to 6S RNA and nucleolar proteins give a nucleolar pattern.
In the last few decades, many nuclear antigens that react with connective tissue disease sera have been described
and characterized. Along with the description of new ANA has come differing and overlapping nomenclature for
the respective nuclear antigens. Generally, the nuclear antigens may be divided into three categories: the nucleic
acids (DNA and RNA), the histone (basic) proteins, and the nonhistone (acidic) proteins. Detailed characterisation
of the autoantibodies is important since different autoantibodies can be associated with specific diseases.
Specific autoantibodies detected by the use of Hep2000 cells include:
Homogeneous Indicates presence of antibodies to dsDNA, ssDNA or histones. Suggestive of SLE but also found
in other CTDs, drug-induced lupus and rarely CAH.
Speckled Caused mainly by antibodies to Sm, RNP, Ro and La (see anti-ENA below). Pattern found in SLE,
MCTD and SS but also other CTDs.
Nucleolar High titre associated with scleroderma, low titre found in other CTDs.
Centromere Most frequently found in CREST (Calcinosis, Raynaud's phenomenon, Oesophageal dysmotility,
Sclerodactyly and Telangiectasia) variant of Progressive Systemic Sclerosis but alos occasionally in other CTDs.
Homogeneous/ Caused by antibodies to Scl-70. These antibodies are very specific to scleroderma and are
Nucleolar frequently associated with the more severe diffuse form of the disease.
Speckled/ Caused by antibodies to La which are found predominantly in SLE and SS
Nucleolar
PCNA These are found in 4% of patients with SLE, but they are not found in patients with rheumatoid arthritis,
scleroderma or polymyositis.
Mitotic Spindle Found rarely in a variety of CTDs
Centriole Found rarely in a variety of CTDs
Nuclear Dots Strongly associated with PBC but also found in CAH and sicca.
Antibodies to Double-Stranded DNA (anti-dsDNA)
Introduction
Antibodies to DNA were initially detected in sera of patients with systemic lupus erythematosus (SLE), and their
unique relationship to this disease was immediately apparent. However, it has also become evident that DNA is a
molecule with multiple epitopes and that antibodies to DNA may include a heterogeneous group of
immunoglobulins with a variety of specificities. Probably most common among the various antibodies reactive
with DNA are those directed against antigenic determinants found on single-stranded DNA (purine and pyrimidine
nucleotide determinants). Detection of such antibodies has little diagnostic specificity; they are found in a wide
variety of autoimmune and connective tissue diseases. In contrast, antibodies reactive primarily or exclusively
with native, double-stranded DNA (herein referred to as anti-dsDNA) show strong association with SLE. These
antibodies are probably directed against deoxyribose-phosphate determinants. Numerous clinical investigations
have confirmed both the relative diagnostic specificity of these antibodies for SLE and the consistent correlation of
their levels with disease activity in many patients.
The practical utility of measuring anti-DNA has led to the development of a number of techniques for their
quantitation. We use an anti-dsDNA ELISA which is more sensitive but the increased sensitivity does appear to
mean that the antibodies are detected in a wider range of diseases.
Use of Test
An anti-dsDNA test is often useful diagnostically for patients who have antinuclear antibodies or clinical findings
suggestive of SLE. The presence of anti-dsDNA is one of the American Rheumatism Association criteria for SLE
and the antibody is rarely found in high levels in patients with other connective tissue diseases. However,
anti-dsDNA antibodies are frequently found in patients with overlap symptoms between SLE and other
autoimmune diseases.
Method Used
Two methods are used:
1. An ELISA for IgG anti-dsDNA.
2. IIF using fixed Crithidia luciliae as substrate.
Reporting of Results
ELISA: The results are reported as IU/ml (using the W.H.O. standard WO80). The turnaround time is 7 working
days.
IFF: The results are reported as „positive‟, „borderline‟ or „negative‟.
Interpretation of Results
The absence of anti-dsDNA antibodies by ELISA is strongly indicative of a disease other than SLE. Antibodies
to dsDNA are not usually found in cases of drug induced lupus.
In SLE actual levels do not correlate well with disease activity though rapid rises in antibody levels do suggest
increases in disease activity.
Antibodies to Extractable Nuclear Antigens (anti-ENA)
Introduction
Different systemic connective tissue diseases have distinct ANA profiles. Thus, the ANA profile can be helpful in
the differential diagnosis of patients with systemic lupus erythematosus (SLE), mixed connective tissue disease
(MCTD), scleroderma, Sjögren's syndrome (SS), rheumatoid arthritis (RA) and other rarer CTDs..
In addition to determining ANA by IIF as described above the measurement of a group of antibodies known
collected as anti-ENAs has recently become routine. Most of these were initially identified by
counterimmunoelectrophoresis and immunodiffusion but they are now usually measured by ELISA. In this
laboratory antibodies to Sm, RNP, Ro, La, Scl-70 & Jo-1 are measured.
Use of Test
Assays to determine ENA specificities are indicated in the evaluation of any patient suspected of having a systemic
connective tissue disease. In most cases ENA determination is indicated only when the ANA is positive.
However, there is a group of patients who have mainly skin involvement SLE, but who have a negative ANA. They
have been found to be positive for antibody to SS-A/Ro antigen.
Method Used
Both the Screen and Profile are performed using ELISA for IgG autoantibodies..
Reporting of Results
ENA Screens are reported as „Negative‟, „Borderline‟ or „Positive‟. ENA profile results are reported in U/ml
with >25 is positive.
Interpretation of Results
The clinical interpretation of the ELISA assays depends on the ENA specificities that are detected.
Antibodies to Sm and RNP Antigens
Antibody to Sm antigen is found in approximately 25% of patients with SLE. There is considerable disagreement
about incidence of this antibody (there appear to be significant racial differences and reports differ on whether or
not it varies with the progress of the disease) but all are agreed that it is strongly associated with SLE. The
presence of antibody to RNP is found in patients with a variety of systemic connective tissue diseases, including
SLE, discoid LE, RA, and SS. It is frequently associated with antibodies to Sm. However, if a high titre of RNP is
found and other ANA specificities are absent, this ANA profile strongly suggests MCTD even though the full
range of clinical involvement is not apparent at that time.
Antibodies to Ro (SS-A) and La (SS-B) Antigens
There are several clinical associations between anti-Ro antibody and certain systemic connective tissue diseases.
This antibody is found in 60 to 70% of patients with Primary Sjögrens syndrome and 30 to 40% of patients with
SLE. There seems to be a close association between the presence of this antibody system and photosensitivity.
Antibody to La antigen is detected in approximately 15% of SLE (70-85% SLE sicca) and 5% RA sicca sera. It is
detected in a higher percentage (60-70%) of sera of patients with Primary Sjögrens syndrome often in the absence
of Ro. Except for the overlapping presence of SS in patients with SLE, there do not appear to be any
distinguishing clinical features which are associated with the presence of anti-La. Ro and La are frequently
detected together.
The screening of pregnant lupus patients for anti-Ro can be important as it is associated with congenital heart block
in the fetus from 18-20 weeks gestation.
Antibodies to Scl-70
Anti-Scl-70 identifies DNA Topoisomerase-1 and is one of several antibodies which give rise to nucleolar staining.
This antibody is associated with the more diffuse form of scleroderma and with impaired pulmonary diffusion. It
also identifies a subgroup of SLE with pulmonary hypertension and nephritis.
Antibodies to Jo-1
Anti-Jo-1 is the most common of a group of myositis-specific autoantibodies (MSAs) which includes anti-PL-12,
anti-OJ, anti-Mi-2 and anti-SRP. The antibody targets are all aminoacyl-tRNA synthetases (histidyl-in the case of
Jo-1). Jo-1 occurs in polymysositis and dermatomyosistis and is usually associated with interstitial lung disease.
A S D MP P D R R
n L r C r r e h A
t E u T i o r e
i g D m g m u p
b i a r a m l
o n r e t a u
d d y s o t s
i u S s / o
e c j i i S
s e ö v P d j
d g e o ö
t r l A g
o L e S y r r
E n y m t e
s s y h n
t o r s
S e s i
y m i t
n i t i
d c i s
r s
o S
m c
e l
e
r
o
s
i
s
N 9 9 1 7 5 3 2 3
u 0 5 0 0 5 0 5 0
c - 0 - - -
SU l 9 7 4 3
M e 5 5 0 0
M i
A i
R d 6 - - - - - 1
Y s 0 0
O D -
Pe
F N 8
rc
A A 0
en
N s 6 1 1 1 1 3
ta
TI s 0 5 0 1 0 0 5
ge
-N D - - - 0 - - -
of
U N 7 3 2 - 2 2 4
Se
C A 0 0 0 3 0 0 0
ra
L 0
Po
E
siti
A H 4 9 - - - - 1
ve
R i 0 5 5
for
A s - - -
Au
N t 7 1 2
to
TI o 0 0 0
an
B n 0
tib
O e
od
D s
ies
Y R 3 - - 6 - - - 9
P o 0 0
R - -
O 5 7
FI 0 0
L 1
E
MISCELLANEOUS AUTOANTIBODIES
Anti-Cardiolipin Antibodies (ACA)
Introduction
Anti-cardiolipin antibodies are one of a group of anti-phospholipid antibodies. There is considerable overlap
between these autoantibodies and lupus anticoagulant. They are found in approx 50% of patients with SLE in
which they are associated with venous and arterial thrombosis. They are also associated with anti-phospholipid
antibody sysndrome in which there is arterial or venous thrombosis, recurrent foetal loss, thrombocytopoenia and
neurological disorders. Anti-cardiolipin antibodies are the anti-M1 anti-mitochondrial antibodies and are
responsible for the false positive VDRL.
Use of Test
Determination of possible cause for myocardial infarct, stroke, peripheral arterial and venous thrombosis
especially in patients under 50 years of age. May contribute to spontaneous abortions especially around 10 weeks
duration and is a cause of neurological events especially in cerebral lupus.
Method Used
ELISA for IgG ACA
Reporting of Results
Results are reported as U/ml where > 10 IU/ml is positive.
Interpretation of Results
Positive results correlate with a predisposition for thrombosis, foetal loss, and thrombocytopoenia in patients with
SLE but levels do not correlate well with disease activity.
Anti-cardiolipin antibodies, the lupus anticoagulant test and the VDRL detect overlapping but not identical
populations of antibodies. Therefore both anti-cardiolipin antibodies and lupus anticoagulant activity should be
measured.
Low levels are also found in infections and some vasculitides.
Rheumatoid Factors (RF)
Introduction
Rheumatoid factors are autoantibodies of IgM, IgG, IgA or even IgE class which recognise an antigenic
determinant on the Fc region of IgG. The exact nature of the determinant is a subject of some debate. Since the
rheumatoid factor is detected in the presence of a vast excess of IgG in the serum, the antigen which is detected is
often referred to as altered IgG. An alternative explanation is that the anti-IgG antibodies are of low affinity and
are only detected when the IgG is aggregated upon coating plates or latex particles.
Although high levels of rheumatoid factors are frequently found in rheumatoid arthritis especially in cases with
extra- articular diseases, they are present, sometimes in even higher levels in a number of other diseases such as
Sjögrens syndrome or Bacterial endocarditis.
Use of Test
Diagnosis of rheumatoid arthritis, differential diagnosis of rheumatic disease.
Method Used
ELISA for IgM Rheumatoid Factor
Reporting of Results
Positive results are those greater than the Reference Range quoted in the Test Report
IgM RF are frequently found in the absence of other classes. It is very rare to detect IgA or IgG RF in the
absence of IgM RF ie in the absence of a positive RA latex test. However, very high levels of IgA or IgG RF can
be detected in the presence of moderate levels of IgM RF.
Interpretation of Results
Raised RF values are very occasionally found in healthy individuals but low levels are common in a wide range of
autoimmune or infectious diseases especially those associated with hypergammaglobulinaemia including viral
hepatitis, chronic liver disease, syphilis, sarcoidosis, leprosy, pulmonary fibrosis. Healthy people over 75
frequently have elevated RF.
High levels are most common in Rheumatoid Arthritis, Subacute Bacterial Endocarditis (SBE) and Sjögrens
syndrome. A very high level of all classes of RF in RA is often associated with extra articular activity although
the titres do not correlate well with disease activity. RF are of little use in monitoring disease activity in RA
where CRP levels should be used.
Anti-Citrullinated Cyclic Peptide (Anti-CCP)
Anti-CCP kits are currently being evaluated with the intention of replacing RF testing by Summer/2008 or as
soon as possible thereafter.
Anti-Acetylcholine Receptor Antibodies (AChRAb)**
Introduction
Antibodies to the acetylcholine receptor are almost invariably associated with myasthenia gravis. The receptor is
a complex of proteins found in skeletal muscle motor endplates. There are several kinds of antibodies to AchR,
ie, some that react to determinants other than the neurotoxin-binding site, others that react with the
neurotoxin-binding site or the extrajunctional receptors, and others that are species specific. The antibodies are
IgG and are pathogenic.
Use of Test
Diagnosis of myasthaenia gravis.
Interpretation of Results
Anti-acetylcholine receptor antibodies are found in 90% of patients with active adult onset generalised myasthenia
gravis. The correlation between the concentration of receptor antibody and the severity of clinical symptoms
appears to be rather low. However, the correlation is much higher in the individual patient, where changes in
symptoms and signs very often coincide with changes in the concentration of antibodies to AChRAb These
autoantibodies are only present in 50% of patients with ocular myasthesia alone.
The test should be used for diagnosis only. Follow up is according to severity of symptoms.
Myasthenic syndromes of childhood seldom show anti-acetylcholine receptor antibodies. The antibodies are
invariably absent in polymyositis.
Anti-Hu , anti-Yo & anti-Ri Antibodies**
These are found in paraneoplastic neurological syndromes and are detected using sections of monkey cerebellum.
Anti-Hu antibodies are associated with encephalomyelitis, sensory neuropathy and rarely autonomic neuropathies
with gastrointestinal dysmotility.
Small cell lung carcinomas are found in 80% of cases.
Anti-Yo antibodies are associated with subacute cerebellar ataxia. 90% of cases have carcinoma of ovary or
breast.
Anti-Voltage Gated Calcium & Potassium Channel Antibodies**
Anti-calcium channel antibodies are found in the paraneoplastic Lambert-Eaton syndrome anti-anti-potassium
channel antibodies are found in Isaac‟s neuromyotonia syndrome..
Anti-Glutamic Acid Decarboxylase Antibodies (Anti-GAD)**
These can be found in patients with “Stiff-Man Syndrome” in approximately 40 % of cases as well as a majority
of patients with newly diagnosed IDDM.
Anti-Ganglioside Antibodies**
These are often found in a variety of autoimmune peripheralnervous system neuropathies such as multifocal
motor neuropathy, Guillain-Barré and Miller-Fisher syndrome.
Anti-MuSK Antibodies**
Antibodies to muscle-specific kinase are often found together with AChRAb in myasthenia gravis
(MG) but also in a majority of patients with AChRAb-negative MG. They are strongly associated
with the ocular form of MG.
Anti-Pituitary Antibodies**
There are a variety of anti-pituitary antibodies associated with such disorders as idiopathic
panhypopituitarism, GH deficiency, ACTH deficiency and acromegaly.
Anti-Parathyroid Antibodies*
These are found found in hypoparathyroidism and polyglandular endrocine syndrome, type 1
Anti-Histone Antibodies*
External Referral Laboratories
* Mr Kevin Green,
Laboratory Manager,
Supraregional Protein reference Unit,
Department of Immunology,
PO Box 894,
Sheffield, S5 7YT
** Neurosciences Group,
Institute of Molecular Medicine,
John Radcliffe Hospital,
Oxford OX3 9DU
The results of all these tests that are sent to specialised laboratories are reported directly to the requesting
physicians.
It should be noted that this whole process is very expensive and time consuming .
GUIDELINES FOR REPEAT TESTING
A S D MP P D R R
n L r C r r e h A
t E u T i o r e
i g D m g m u p
b i a r a m l
o n r e t a u
d d y s o t s
i u S s / o
e c j i i S
s e ö v P d j
d g e o ö
t r l A g
o L e S y r r
E n y m t e
s s y h n
t o r s
S e s i
y m i t
n i t i
d c i s
r s
o S
m c
e l
e
r
o
s
i
s
N 9 9 1 7 5 3 2 3
u 0 5 0 0 5 0 5 0
c - 0 - - -
l 9 7 4 3
e 5 5 0 0
i
i
d 6 - - - - - 1
s 0 0
D -
N 8
A 0
s 6 1 1 1 1 3
s 0 5 0 1 0 0 5
D - - - 0 - - - ANA Q&A
N 7 3 2 - 2 2 4
A 0 0 0 3 0 0 0
0
H 4 9 - - - - 1
i 0 5 5
s - - -
t 7 1 2
o 0 0 0
n 0
Q. What does the test detect?
A. Serum antibodies (usually IgG) to antigens on or inside the nucleus
Q. How is the test performed?
A. Usually by immunofluorescence on fixed cells. These days the cell of choice is almost always HEp2 (a
human epithelioma cell line) which has very large nuclei enabling many ANA subtypes to be identified.
Q. How should the blood sample be collected?
A. The sample should be clotted so that serum can be used for the test. Fibrinogen may interfere with the test.
Q. When should an ANA be requested?
A. Any patient suspected of having a connective tissue disease or any autoimmune disease.
Q. What is the significance of a positive result?
A. This will depend very much on the subtype. The most commonly seen patterns are:
1. Homogeneous. Strongly associated with
2. Speckled. Found most frequently in SLE, Sjogren‟s and
SLE
MCTD
3. Centromere. Strongly associated with CREST
syndrome
4. Nucleolar. Frequently founded in
scleroderma
5. Nuclear Dots. This is a rare pattern but if found in PBC suggests a poor
prognosis
Q. When should a patient be referred to a specialist?
A. All positive samples are titred and a titre of 160 or greater is considered positive. It should be remembered,
however, that the incidence of ANAs increases markedly with age and that the test is only an aid to diagnosis.
Q. What are the pitfalls of ANA testing?
A. Methodological: There are many antibodies that can give rise to a similar ANA pattern. eg. antibodies to
Sm, RNP and La will all give a speckled pattern but are associated (but not diagnostic) for different connective
tissue diseases (SLE, MCTD and Sjogren‟s in this case). A speckled ANA should always be further subtyped.
Diagnostic: These antibodies are associated with a wide variety of connective tissue diseases but are
also found in other diseases and in apparently normal people. There presence must therefore always be
interpreted in the light of a full diagnostic investigation.
MEASUREMENT OF COMPLEMENT PROTEINS
Introduction
Serum complement components are measured for the detection of:
1 Inherited deficiencies of complement components (which are very rare)
2 Complement activation during infectious diseases or immune complex diseases (which is much more common)
Complement deposition in tissues is also detected in biopsy material (skin, kidney) to diagnose immune complex
disease.
Inherited Deficiencies
Genetic deficiencies of almost all of the complement components have been detected very rarely in individuals.
Deficiency of C3 and control proteins of the alternative pathway which lead to acquired deficiency of C3 are
associated with recurrent bacterial infection with a variety of organisms. Deficiencies of the late components are
also associated with recurrent infections, usually Neisserial. Deficiencies of the classical pathway are more
commonly associated with immune complex disease rather than infection, demonstrating the key role of
complement in the handling of immune complexes. Deficiency of the control protein C1-inhibitor is the cause of
a unique disease, hereditary angioedema, which is an autosomal dominant trait characterised by intermittent
swelling of limbs and internal viscera.
Disease Associations of Complement Deficiencies
A S D MP P D R R
n L r C r r e h A
t E u T i o r e
i g D m g m u p
b i a r a m l
o n r e t a u
d d y s o t s
i u S s / o
e c j i i S
s e ö v P d j
d g e o ö
t r l A g
o L e S y r r
E n y m t e
s s y h n
t o r s
S e s i
y m i t
n i t i
d c i s
*C4 levels are the result of two genes C4A and C4B, both show codominant expression of alleles. 15% of the
r
population have lack of s of the four alleles. Some are associated with SLE.
one
o S
m c
e l
e
r
o
s
i
s
N 9 9 1 7 5 3 2 3
u 0 5 0 0 5 0 5 0
c - 0 - - -
l 9 7 4 3
e 5 5 0 0
i
i
d 6 - - - - - 1
Disease Associations of Acquired Changes in Levels of Complement Components
A S D MP P D R R
n L r C r r e h A
t E u T i o r e
i g D m g m u p
b i a r a m l
o n r e t a u
d d y s o t s
i u S s / o
e c j i i S
s e ö v P d j
d g e o ö
t r l A g
o L e S y r r
E n y m t e
s s y h n
t o r s
S e s i
y m i t
n i t i
d c i s
r s
o S
m c
e l
e
r
o
s
i
s
N 9 9 1 7 5 3 2 3
u 0 5 0 0 5 0 5 0
c C4 0
C3 and - levels are- routinely measured and C1-Inhibitor levels in suspected cases of deficiency. Other
- -
components are not routinely measured.
l 9 7 4 3
e 5 5 0 0
Determination of C4
i
i
Introduction - - - - 1
d 6 -
s 0 0
a protein of the classical pathway of complement. Activation of this pathway occurs predominantly
C4 is D -
because of the presence of deposited IgG or IgM containing immune complexes. The products of C4 activation
N 8
are rapidly removed from the circulation which results in lowered C4 levels. Depletion of C4 is thus frequently
A 0
s 6 1 of
an indication 1 immune 3complex disease. One complication is that C4 is an acute phase protein whose
1 1
s 0 5 0 0 0 5
concentration will1rise during the acute phase of an infective or autoimmune disease.
D - - - 0 - - -
N 7
Use of Test3 2 - 2 2 4
A 0 0 0 3 0 0 0
0
Diagnosis and monitoring of immune complex disease eg SLE, rheumatoid arthritis or immune mediated
vasculitis.
H 4 9 - - - - 1
i 0 5 5
Diagnosis of hereditary angioedema or other C1-inhibitor deficiencies. C1 inhibitor deficiency does not occur in
s - of -
the absence - a low C4 level.
t 7 1 2
o Used
Method 0 0 0
n 0
e
Nephelometry
s
R 3 - - 6 -
Reporting of Results - - 9
o 0 0
- -
5 7
0 0
1
º
2
Clinically significant results are those 100 0 kUA/1 Extremely High Level
The turnaround time for these tests is 10 working days.
Total IgE Reference Range
18 years 0-250 KU/l
Determination of Tryptase Activity
Activated human mast cells secrete preformed, granule derived, mediators including histamine, proteoglycans and
the neutral preteases, tryptase and chymase, together with newly formed mediators. The later include
prostglandins, leucotrienes and platelet activating factor. Since tryptase is specific marker of the secretory
granule, its presence in various body fluids is a reflection of mast cell degranulation.
Tryptase is a serine esterase and basal concentrations are in the order of 0.2 - 8 ug/L with concentrations of more
than 13.5 ug/L following allergic stimulation or mast cell degranulation. Levels as high as 100 ug/L may be seen
in the extreme clinical situations, such as a severe reaction to anaesthetic agents.
Tryptase is catabolised by the liver with an in vivo half-life of 3 hours compared to the 2-3 minutes of histamine.
They decay curve after allergic stimulation and mast cell degranulation is observed over 24 hours compared with
the 20 minutes for histamine. Tryptase is stable in isolated plasma or serum.
Isolated tryptase concentrations may be of value in the assessment of allergic disorders and mast cell syndromes
e.g. mastocytosis.
Investigation of anaphylactic reactions would be better conducted with serial samples collected within one hour
of reaction and subsequently at 3, 8 and 24 hours, or at similar intervals, if it is not possible to collect the samples
at these times. Note that it is essential that all samples and accompanying Request Forms state the time of the
adverse reaction and also have the precise time and date of the sample so that the results can be interpreted in
relation to the event thought to trigger off mast cell degranulation.
The turnaround time is 5 working days.
SPECIFIC IgG ANTIBODY TEST AGAINST FUNGAL & AVIAN PRECIPITINS
IgG antibodies are part of the natural defence system in the body and are produced in response to contact with
foreign substances. If intense exposure to these antigens continues over a long period of time, clinical disease
may develop in some people.
Clinical symptoms of Extrinsic Allergic Alveolitis (EAA), Farmer's Lung or Bird Fancier's Lung may include
shortness of breath, aching joints and muscles and a general feeling of being unwell.
EAA can occur alone or with Allergic Broncho- Pulmonary Alveolitis (ABPA). If left untreated ABPA can lead
to permanent lung damage (fibrosis). Aspergilloma, a fungus ball, may develop with chronic lung disease.
Although the incidence of ABPA is low, patients with asthma or cystic fibrosis are more susceptible to this
disease and early diagnosis is essential to limit lung damage.
Traditionally, the Ouchterlony immunodiffusion method has been used to demonstrate qualitatively, the presence
of precipitin antibody to a specific antigen. These tests are technically demanding to perform and only
semi-quantitative
The automated test that we have adopted (solid phase immunoassay) provides a quantitative result (in mg/l)
which can be used to monitor patients and the clinical effectiveness of treatment over a period of time
Measuring Range
The measuring range for specific IgG antibodies is 2-200mg/l
Quantitative tests for IgG antibodies against the following antigens are available
Gm3 Aspergillus fumigatus
Gmx7 Combination of Micropolyspora faeni & Thermoactinomyces vulgaris
Ge91 Pigeon serum proteins, feathers & droppings
Ge90 Budgerigar serum proteins, feathers & droppings
Ge92 Parrot serum proteins, feathers & droppings
Note – this is not an exhaustive list, please contact the Immunology Laboratory for additional tests.
Interpretation of Specific IgG Antibody levels
Aspergillus precipitins
0 - 39 mg/l Unlikely to indicate aspergillosis
40 – 89 mg/l Likely to indicate aspergillosis if patient has appropriate clinical features
90 mg/l or greater Highly likely to indicate aspergillosis
Note: In non-CF patients 40 is the cutoff whilst in CF patients 90 is the cut off
Farmers lung (Micropolyspora faeni & Thermactinomyces vulgaris precipitins)
0 – 39 mg/l Unlikely to indicate Farmers lung
40 - 89 mg/l Likely to indicate Farmers lung if patient has appropriate clinical features
90 mg/l or greater Highly likely to indicate Farmers lung
Avian precipitins (Pigeon, Parrot and Budgerigar)
0 - 9.9 mg/l Unlikely to indicate significant reaction to Bird Antigens
10 mg/l or greater Likely to indicate significant reaction to Bird Antigens