Immunology Services

We offer a range of tests to facilitate the diagnosis of autoimmune and neuroimmunological disease, blood cancers, primary and secondary immunodeficiency, infections and allergy.

Available Assays and Reference Ranges

Autoimmunity and Neuroimmunology

Adrenal cortical antibodies

(Serum: Negative)
(Up to 14 days)

Test for autoimmune adrenal disease.

Also see endocrine antibodies


Anti-nuclear antibodies

(Serum: Titre <1:100)

Low ANA titres of 1:100 (positive fluorescence at serum dilution of one in one hundred) are generally not significant in adults, but can be in children.

(Up to 4 days)

ANA’s are associated with a variety of conditions other than SLE including rheumatoid diseases, chronic active hepatitis, fibrosing alveolitis, viral infections and drug ingestion. Patterns of ANA are said to be significant: Nucleolar associated with scleroderma, centromere with CREST syndrome, and speckled pattern with MCTD, Sjögrens, SLE and Polymyositis. Rim or homogeneous has been associated with SLE but there is a considerable amount of pattern overlap. High titre ANA at 1:1600 are strongly suggestive of connective tissue disease.


Anti-C1q autoantibodies

(Serum: 0 – 20 units/ml)*
(Up to 28 days)

Autoantibodies against C1q are a major criterion in the diagnosis of hypocomplementaemic urticarial vasculitis. They are also found in up to 50% of SLE patients and 95% of patients with lupus nephritis. C1q antibodies may be useful for assessing the risk of renal flares, and also for monitoring the effectiveness of immunosuppressive treatment in active lupus nephritis.


Aquaporin 4 antibodies

(Up to 14 days)

See NMO antibodies


B2GP1 antibodies

(Serum: 0 – 20 U/ml)*
(Up to 7 days)
[Also see cardiolipin antibodies]

B2GP1 is a 50kD plasma protein (apolipoprotein H) that inhibits the intrinsic coagulation pathway, ADP mediated platelet aggregation and the prothrombinase activity of activated platelets.  “Anti cardiolipin antibodies” bind to an altered form of B2GP1 which may be reproduced by binding B2GP1 directly to an ‘ELISA’ plate. The detection of anti-B2GP1 antibodies is said to have enhanced specificity for Anti-phospholipid syndrome (APS) and related coagulation disorders over the traditional anti-cardiolipin assay, which may display some false positive results due to cross reactivity of these antibodies with some infectious disease related antigens.  This is currently a quantitative IgG antibody assay.


Cardiac antibodies

(Serum: Negative)
(Up to 14 days)

Cardiac antibodies(Serum: Negative)(Up to 14 days) Though the diagnostic value is low these antibodies are found in some patients with Dressler’s syndrome, following myocardial infarction, after cardiac surgery and in some cardiomyopathies.


Cardiolipin/Phospholipid antibodies

(Serum: IgG: 0 – 15 GPLU/ml
IgM: 0 – 12.5 MPLU/ml)*
(Up to 7 days)
[Also see B2GP1 antibodies]

Antibodies have been associated with SLE, recurrent miscarriages and arterial and venous thrombosis. Slightly elevated levels may be found in some infections and so only positive results at two time points at least 6 weeks apart are considered significant. IgG and IgM antibodies are assayed separately. Significant levels of antibodies do not necessarily correlate with the severity of the disease.

Please note that lupus anticoagulant is performed in haematology.


CSF Tau protein:asialo-transferrin

Present only in CSF
(Up to 4 days)

Ideal suspected CSF fluid volume = 250 microlitres, but not less than 50

Cerebrospinal rhinorrhoea is potentially serious due to risk from infection. In patients presenting with a nasal discharge of clear fluid it is important to identify the nature of the fluid.  CSF is readily identified by the presence of asialo-transferrin (Tau protein). This laboratory offers a reliable, sensitive and simple electrophoretic method for the rapid identification of Tau protein.


Cyclic citrullinated peptide (CCP) antibodies

(Serum: < 7 U/ml)*
(Up to 4 days)

Anti-CCP antibodies are potentially important surrogate markers for diagnosis and prognosis in rheumatoid arthritis (RA), because they:

  • are as sensitive as, and more specific than, IgM rheumatoid factors (RF) in early and fully established disease
  • may predict the eventual development into RA when found in undifferentiated arthritis
  • are a marker of erosive disease in RA
  • may be detected in healthy individuals years before onset of clinical RA

dsDNA antibodies

(Serum: EIA: < 30 IU/ml *
Crithidia IIF: Negative)
(Up to 7 days)

Assay of antibodies to native, double stranded DNA (dsDNA antibodies), is carried out on all patients with SLE, as a qualitative test by IIF on the kinetoplast of crithidia lucillae which is then followed up with a quantitative assay by EIA. dsDNA antibodies may be detected in the absence of ANA and are extremely useful in monitoring the activity of the disease.


Endocrine abs (Adrenal, Ovary, Testis)

(Serum: Negative)
(Up to 14 days)

Adrenal antibodies can be associated with autoimmune Addison’s disease where a gradual destruction of the adrenal gland leads to adrenocortical insufficiency.  Steroid cell antibodies can also be associated with premature ovarian failure and premature testicular failure. They are also associated with the autoimmune polyglandular syndrome types 1,2 and 3.

Autoimmune endocrinopathy may be seronegative in a minority of cases.


Endomysial abs

(Serum: Negative)
(Up to 14 days)

IgA abs directed against theendomysium are detected in 70% of patients with dermatitis herpetiformis and >90% of patients with untreated coeliac disease but are rarely present in normal individuals or in patients with other enteropathies.  Decreasing antibody titres correlate well with adherence to gluten free diet.


Extractable Nuclear Antigen (ENA) antibodies

(Serum: 0 – 20 EU/ml)*
(Up to 7 days)

ENA antibodies recognise saline extracted nuclear antigens. There are many specificities recognised of which this laboratory currently offers six:

  • Sm (a marker for SLE);
  • RNP (said to be present in >95% MCTD);
  • SSA [Ro] (associated with cutaneous lupus, SLE, neonatal lupus & congenital heart block);
  • SSB [La] (SLE, Sjögrens syndrome);
  • Jo1 (30% of polymyositis cases) and
  • Scl70 (associated with systemic sclerosis).

Patients with SLE or Sjögrens should be screened for ENA antibodies especially females considering pregnancy.


Ganglioside antibodies GD1b

(Serum: IgG  <1:500
IgM <1:500)§

(Up to 14 days)

Antibody to the ganglioside GD1b has been associated with motor or sensorimotor neuropathies.  High titres of anti GM1 are most typical of multifocal motor neuropathy but antibodies to other gangliosides such as GD1b and asialoGM1 may also be detected. Low titres of antibodies directed against GD1b, GM1 and asialoGM1 may also be detected in amyotrophic lateral sclerosis and Guillain-Barré syndrome.  These antibodies are currently screened in house and positives are sent to Neurology, Southern General Hospital, Glasgow, for quantitation.


Gangliside antibodies GM1

(Serum: IgG <1:500
IgM <1:500
§
(Up to 14 days)

The presence of antibodies directed against GM1 (monosialoganglioside GM) has been associated with motor and sensorimotor neuropathies and in particular with multi- focal motor neuropathies. Lower titre of GM1 antibodies may also be found in amyotrophic lateral sclerosis and Guillain - Barré syndrome. a’GM1 antibodies may occur as either polyclonal or IgM monoclonal antibodies. The carbohydrate moiety of GM1, in particular the galactose and sialic acid residues, is the site of antibody binding to gangliosides. Due to the presence of similar moieties on other gangliosides low levels of antibody cross-reaction may be experienced in tests for gangliosides other than GM1.  These antibodies are currently screened in house and positives are sent to Neurology, Southern General Hospital, Glasgow, for quantitation.


Ganglioside antibodies GQ1b (Miller Fisher syndrome)

(Serum: <1:500 (IgG & IgM))§
(Up to 14 days)

These antibodies are currently screened in house and positives are sent to Neurology, Southern General Hospital, Glasgow, for quantitation.


Ganglioside antibodies sulphatide (Sensory neuropathy)

(Serum: <1:10000 (IgG & IgM))§
(Up to 14 days)

These antibodies are currently screened in house and positives are sent to Neurology, Southern General Hospital, Glasgow, for quantitation.


Gastric parietal cell antibodies

(Serum: Negative)
(Up to 4 days)

Intrinsic factor antibodies should be carried out in conjunction with GPC antibodies

These antibodies are present in up to 90% of patients with atrophic gastritis and pernicious anaemia. Also present in gastritis without anaemia (12%), autoimmune thyroid disease (30%), Addison’s disease (25%) and iron deficiency anaemia (20%).


Gliadin deamidated peptide (DP) antibodies

(Serum: IgG < 7 U/ml)*
(Up to 7 days)

IgA anti-tissue transglutaminase (tTG), antibodies are specific for coeliac disease, but can be negative in patients with IgA deficiency. In this situation IgG anti-gliadin DP antibodies can be clinically useful. The titre of these antibodies decreases with gluten free diet, as does the level of endomysial antibodies and tTG antibodies.

It has been reported that IgA deficient patients have a ten to fifteen fold increased incidence of coeliac disease. It is therefore suggested that IgG anti-gliadin DP antibodies are carried out in all IgA deficient individuals.


Glomerular basement membrane (GBM) antibodies

(Serum: < 7 U/ml)*
(Up to 4 days)

Test for Goodpastures syndrome. Antibodies to the non-collagenous portion of type IV collagen are detected by ELISA method as indirect immunofluorescence is both less sensitive and less specific being positive in only 75%, or less, of proven cases. Urgent requests for GBM antibodies (as with ANA, ANCA and dsDNA antibodies) must be arranged with the laboratory.


Glutamic acid decarboxylase antibodies: Stiff Man syndrome

(Serum: 0 - 10 IU/ml)*
(Up to 14 days)

GAD index

GAD Index is available to determine CSF specific GAD synthesis (requires CSF and serum).

Glutamic acid decarboxylase (GAD) is an enzyme concentrated in neurons, which control muscle tone and exteroreceptive spinal reflexes. High levels of antibodies to GAD are found in ~60% of patients with Stiff man syndrome; in IDDM the titres are much lower. The contribution of GAD antibodies to IDDM has not been proved. GAD has also been implicated in autoimmune encephalitis.


Intrinsic factor antibodies

(Serum: < 6 U/ml)*
(Up to 7 days)

Detected in 70% of patients with pernicious anaemia. This test should be carried out together with gastric parietal cell antibodies.


Isoelectric focusing (Oligobanding) IgG

(Paired serum and CSF: Clinical comment is supplied with each report)
(Up to 14 days)

Please enclose CSF protein value with each request

Volume of CSF required: Ideally 1-2ml but minimum of 250 microlitres. Contact the lab if insufficient volume.

Oligobanding refers to discrete populations of immunoglobulin detected by electrophoresis in CSF, which are NOT matched in serum from the same patient. Oligobanding is seen in ~85-95% of patients with clinically proven multiple sclerosis. The assay is useful as a confirmatory test in multiple sclerosis but bands are not specific for this disease as they also occur in cerebrovascular accidents, in infections of the CNS and in pathological processes involving an immune response e.g. encephalitis, neuro-sarcoid and SLE.

Please note that paired samples of CSF and serum are essential for this assay.


Liver antigen antibodies (blot)

(Up to 14 days)

Detection and confirmation of antigen specific antibodies associated with primary biliary cirrhosis and autoimmune hepatitis. These include M2, LKM-1, LC-1, SLA/LP, SP100, GP210 and f-Actin. M2 is also measure quantitatively by ELISA


LKM antibodies

(Serum: Negative)
(Up to 4 days)

These antibodies, which stain the cytoplasm of hepatocytes and proximal renal tubules are found in a subgroup of patients with ANA negative, autoimmune chronic active hepatitis (CAH). LKM1 antibodies are positive in CAH type 2, which is the most common autoimmune liver disease of childhood.


Mitochondrial antibodies

(Serum: Negative)
(Up to 4 days)

Present in >90% of cases of primary biliary cirrhosis, often at high titre (>1:200). Also occasionally present in chronic active hepatitis and halothane induced hepatitis patients but with titres of <1:100. Serum IgM levels are invariably increased.


Mitochondrial (M2) antibodies

(Serum: 0 – 10 EU/ml)
(Up to 7 days)

For those wishing to confirm the presence of mitochondrial antibodies or to monitor patients with a quantitative assay an EIA method is available which distinguishes antibodies to the major enzyme pyruvate dehydrogenase complex (M2) and affords a quantitative assay in EU/ml.


MOG antibodies

(Serum: Negative)
(Up to 14 days)

Another autoimmune cause for neuromyelitis optica is an oligodendropathy attributed to MOG antibody. MOG antibodies target myelin sheath thus causing demyelination.MOG and NMO antibodies are run as a combine test.


Myeloperoxidase (MPO) antibodies

(Serum)
Negative: <3.5 IU/ml*
Equivocal: 3.5 – 5.0 IU/ml*
Positive: >5.0 IU/ml*

(Up to 7 days)

Antibody to myeloperoxidase is associated with organ-limited vasculitis including necrotising and crescentic glomerulonephritis. The assay is useful in confirming MPO specific antibodies in sera which are ANCA-positive. Typically the level of MPO antibodies parallel disease state with increasing levels when vasculitis is active.

Urgent requests must be arranged with the laboratory.


Anti-Neutrophil cytoplasmic antibodies (ANCA)

(Serum: Negative)
(Up to 4 days)

Pattern and titre reported on positives
[See MPO & PR3 antibodies]

c-ANCA is a test for granulomatosis with polyangiitis and microscopic polyarteritis (see also test for proteinase 3). p-ANCA may occur in other vasculitic disorders as well as some forms of glomerulonephritis (see also test for myeloperoxidase).


NMO antibodies

(Serum: Negative)§
(Up to 14 days)

Anti-NMO antibodies are associated with neuromyelitis optica (NMO) also known as Devic’s disease and optic-spinal multiple sclerosis. It is a severe inflammatory demyelinating disease that affects optic nerves and spinal cord without affecting the brain. Aquaporin 4 has been identified a major NMO antigen and the test offer is against this antigen. This test is used to distinguish NMO from multiple sclerosis.

NMO and MOG antibodies are run as a combined test.


Pancreatic islet cell antibodies

(Serum: Negative)
(Up to 14 days)

At the time of diagnosis 75% of type I diabetics have detectable levels of circulating islet cell antibodies. Such antibodies decrease and eventually disappear with duration of disease. Some studies have indicated persistent levels of antibodies in association with polyendocrine disease (type Ib). There have been no reports of antibodies to pancreatic islet cells in type II diabetics.


MAG antibodies

Paraprotein neuropathies

(Serum: Negative)*
(Up to 14 days)

Positive samples are sent away for quantitation by ELISA

Myelin associated glycoprotein (MAG) is a glycoprotein component of the myelin of central and peripheral nervous systems. Monoclonal reactivities against MAG are detected in about 50-75% of patients with IgM paraproteinaemia and peripheral neuropathy. Sera from patients with neuropathy that are negative for MAG antibodies often exhibit reactivity against various gangliosides.


Phospholipase A2 (PLA 2) receptor antibodies

Indirect immunofluorescence

(Serum : Negative)*
(Up to 14 days)

ELISA

(Serum)
Negative: <14 RU/ml*
Borderline: ≥14 – <20 RU/ml*
Positive: ≥20 RU/ml*
(Up to 28 days)

Autoantibodies to the M-type phospholipase A2 receptor (PLA2R) are sensitive and specific for idiopathic membranous nephropathy (IMN), an organ specific autoimmune disease of the glomeruli. The test is helpful both in the diagnosis of IMN and monitoring response to treatment. These antibodies are specific and are found in up to 70% of the patients with IMN.

The ELISA assay will be run on specific request or when the serum sample is “sticky” and prevents processing by IIF


Proteinase 3 (PR3) antibodies

(Serum)
Negative: <2 IU/ml*
Equivocal : 2.0 – 3.0 IU/ml*
Positive : >3.0 IU/ml*
(Up to 7 days)

PR3 antibody is a marker for granulomatosis with polyangiitis and is occasionally detected in microscopic polyarteritis. The quantity of PR3 antibody generally parallels disease activity with higher levels in the active state of the disease. EIA affords a quantitative assay which is useful when monitoring the disease. Antibodies to PR3, are responsible for the characteristic granular cytoplasmic pattern of the neutrophils when stained by IIF.

Urgent requests must be arranged with the laboratory.


Rheumatoid factor

(Serum: 0-14 IU/ml)*
(Up to 4 days)

Also see Cyclic citrullinated peptide (CCP) antibodies

Rheumatoid factors are antibodies which are directed against other immunoglobulins. A latex enhanced turbidimetric assay is used to detect these. Approximately 70% of patients with rheumatoid arthritis are sero positive and antibodies may occur in other conditions including many infections, myeloma, lymphomas, cryoglobulinaemia and connective tissue diseases. Antibodies (RF) may also be found in allegedly normal individuals aged over 75. Titre of rheumatoid factor is less sensitive than sequential assay of CRP when monitoring activity of rheumatoids.


Skin antibodies

(Serum: Negative)
(Up to 14 days)

Antibodies are found in (i) intercellular substance of the epidermis (desmosome), which strongly suggest a diagnosis of pemphigus though these antibodies may also be found in patients with severe burns or a trichophyton infection. (ii) dermal-epidermal basement membrane which is highly specific for bullous pemphigoid and is present in 80% of these patients. A titre is useful in monitoring the disease.


Smooth muscle antibodies

(Serum: Negative)
(Up to 4 days)

Present in high titre in up to 70% of patients with autoimmune hepatitis who may also be positive for mitochondrial, nuclear and dsDNA antibodies (25%)


Striated muscle antibodies

(Serum: Negative)
(Up to 14 days)

In patients with Myasthenia Gravis with thymoma these antibodies are typically positive but in such patients without thymoma the antibodies occur in only 60% of cases. This assay is usually carried out with a test for acetyl choline receptor antibodies but as this latter test is sent away to Oxford for quantitative assay it will not be carried out as a routine unless specifically requested.


Thyroid peroxidise antibodies(microsomal [TPO])

(Serum: < 60 IU/ml)*
(Up to 4 days)

Present at high levels in 95% of patients with Hashimotos thyroiditis, 20% of patients with Graves disease and 90% of patients with primary myxoedema. Antibodies may also be present at low levels in colloidal goitre, thyroid carcinoma, De Quervains thyroiditis, other organ specific auto-immunities and in healthy individuals. If persistent in euthyroid individuals it may indicate autoimmune thyroiditis and predisposition to future thyroid failure.


TSH receptor antibodies

(Serum: Negative)§
(Up to 28 days)

Hyperthyroidism in Grave’s disease is due to autoantibodies to the TSH receptor and measurement of these autoantibodies can be useful in disease diagnosis and management. This assay is currently sent to Immunology PRU, Northern General Hospital, Sheffield.


Tissue Transglutaminase antibodies

(Serum)
Negative: < 7 U/ml*
Equivocal : 7 – 10 U/ml*
Positive : >10 U/ml*
(Up to 4 days)

[Also see gliadin deamidated peptide antibodies]

The endomysial autoantigen has been identified as the protein cross-linking enzyme tissue transglutaminase (tTG). Antigen specific assays provide an alternative to the conventional indirect immunofluorescence assay using primate oesophagus.

We screen for coeliac disease with an IgA anti-TTG assay. A very low result suggests that the patient may be IgA deficient and therefore we proceed to an IgG anti-gliadin deamidated peptide assay which is more sensitive than IgG anti-TTG. We also add immunoglobulins to check for IgA deficiency.

IgA deficiency (partial or complete) is about 1:400 blood donors and 1:40 patients with coeliac disease.

Key for reference ranges

Where a quantitative reference range is provided the origin of that range is as follows:

*

Manufacturers reference range (verified in-house)

National (or international) reference laboratory range (verified in-house)

Internally generated reference range

§

Send away test with accompanying (referral laboratory derived) range

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Blood Cancer

B2 microglobulin (B2M)

(Serum: 0 – 4.0 mg/l)
(Up to 4 days)

Useful for monitoring lymphocyte activation and turnover in myeloma and HIV related diseases. Because B2M is filtered by the glomeruli and metabolised in the renal tubules higher levels are seen in patients with renal dysfunction.


Cryoglobulins

(Serum: Negative)
(Up to 10 days)

Positives may be typed:monoclonal, polyclonal or mixed (IgG/A/M).

When cryoglobulins are associated with myeloma, Waldenströms macroglobulinaemia, or lymphoma they consist of one immunoglobulin isotype but may be mixed or polyclonal in other diseases such as connective tissue diseases. Patients with renal disease and a low C4 level or patients with unexplained cutaneous vasculitis should be screened for presence of circulating cryoglobulin.

Please refer to special conditions of collection and despatch to laboratory (page 6 in the Handbook).


Immunoglobulins (IgG/A/M)

(Serum: IgG: 6.00 – 16.00 g/l 
(adult): IgA: 0.80 – 4.00 g/l
IgM: 0.50 – 2.00 g/l)
(Up to 4 days)

N.B. Reference ranges are age specific and may differ between ethnic groups

Immunoglobulins are an essential request in recurrent infections, lymphoproliferative diseases including myeloma and all cases of ‘failure to thrive’.  IgA deficiency occurs in 1:500 individuals but, transfusion reactions apart, may not be associated with disease. Polyclonal increases of IgG occur in chronic infection and inflammation, chronic liver disease and connective tissue diseases. Raised levels of IgM are found in acute inflammation and in primary biliary cirrhosis. (Markedly elevated IgM in the presence of mitochondrial  antibodies is virtually diagnostic of PBC) Low levels of IgG and IgA may be due to loss (protein losing enteropathy or nephrotic syndrome), reduced synthesis (e.g. lymphoproliferative disorders or primary immunodeficiency) or excessive catabolism. Low immunoglobulins always require further investigation. Where appropriate details are supplied age and sex related normal levels are printed on the report. See also IgG sub-classes and functional antibodies.


IgG Subclasses

(Serum: IgG1: 3.2 – 10.2 g/l
adult: IgG2: 1.2 – 6.6 g/l
IgG3: 0.2 – 1.9 g/l
IgG4: 0.0 – 1.3 g/l)
(Up to 7days)

IgG subclass deficiency is mainly related to IgG1 and IgG2 where individuals may suffer recurrent infections.


Immunoglobulin D (IgD)

(Serum: 0.05-0.20 g/L)
(Up to 14 days)

Serum IgD is measured in the case of IgD myeloma and some forms of periodic fever syndrome.


Immunoglobulin E (IgE)

(Serum: 0 – 90 IU/ml, adult)
(Up to 7 days)

N.B. Reference ranges may differ between ethnic groups

Serum IgE may be helpful in diagnosing atopic diseases however the reference range is very wide and levels do not correlate well with symptoms. Very high levels of IgE are seen both in atopic eczema and in parasitic infestations (especially S Mansoni) and may result in false positive specific IgE to a single allergen.


Isoelectric focusing (Oligobanding) IgG

(Paired serum and CSF: Clinical comment is supplied with each report)
(Up to 14 days)

Please enclose CSF protein value with each request

Volume of CSF required: Ideally 1-2ml but minimum of 250 microlitres. Contact the lab if insufficient volume.

Oligobanding refers to discrete populations of immunoglobulin detected by electrophoresis in CSF, which are NOT matched in serum from the same patient. Oligobanding is seen in ~85-95% of patients with clinically proven multiple sclerosis. The assay is useful as a confirmatory test in multiple sclerosis but bands are not specific for this disease as they also occur in cerebrovascular accidents, in infections of the CNS and in pathological processes involving an immune response e.g. encephalitis, neuro-sarcoid and SLE.

Please note that paired samples of CSF and serum are essential for this assay.


Serum electrophoresis

(Serum: Clinical comment will accompany each report)
(Up to 4 days)

Sera are screened for qualitative abnormalities in proteins especially of the immunoglobulins. Scans demonstrating a monoclonal band are automatically followed up using immunofixation to determine both the isotype and the light chain of the monoclonal protein. Other typical patterns seen on electrophoresis may indicate evidence of acute phase responses, immunodeficiency, etc.

Where myeloma is suspected urine and serum should be sent together.


Serum immunoglobulin free light chains (FLC)

Kappa 3.30 – 19.40 mg/l *
Lambda 5.71 – 26.30 mg/l *
Kappa / Lambda ratio 0.26 – 1.65 *
(Up to 4 days)

This assay may be inaccurate at levels <0.9mg/l. In a small proportion of patients with high serum FLC levels, false negative results may occur as a result of “antigen excess”. Any anomaly between the serum FLC results and other laboratory tests and/or clinical evidence should be reported to the laboratory for re-testing the serum FLC.

Normal plasma cells make more immunoglobulin light than heavy chains and secrete free light chains in amounts detectable in serum (estimated to be 0.5g/day). Serum free light chains are removed by glomerular filtration with a half-life of a few hours. They are not easily detectable in urine until the threshold for tubular reabsorption is exceeded (10 – 20g / day).

Serum FLC measurements are recommended in assessment of all plasma cell dyscrasias and in B cell lymphoproliferative diseases. They are particularly important in diagnosis and management of light chain only myeloma.


Viscosity

(Plasma: 1.50 – 1.72 (c.f. to water))*
(Up to 10 days)

Plasma viscosity is an essential test when monitoring Waldenstroms’ macro-globulinaemia and also when investigating an unexplained retinal or cerebrovascular occlusion. In such patients a cryoglobulin may also be present.

Blood samples can be transported at room temperature but separated plasma should not be refrigerated. Please send EDTA blood.


Key for reference ranges

Where a quantitative reference range is provided the origin of that range is as follows:

*

Manufacturers reference range (verified in-house)

National (or international) reference laboratory range (verified in-house)

Internally generated reference range

§

Send away test with accompanying (referral laboratory derived) range

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Immunity and Infection

Anaphylaxis testing

See mast cell tryptase

Aspergillus - specific IgG antibodies

(Serum:  < 40 mgA/L)*
Also see fungal antigens
(Up to 5 days) 

Specific IgG antibodies directed against aspergillus fumigatus demonstrate previous exposure.

Avian antigens - specific IgG antibodies

(Serum: Negative)
(Up to 5 days)

Specific IgG antibodies directed against budgerigar and pigeon antigens are currently available.


B2microglobulin (B2M)

(Serum: 0 – 4.0 mg/l)
(Up to 4 days)

Useful for monitoring lymphocyte activation and turnover in myeloma and HIV related diseases. Because B2 is filtered by the glomeruli and metabolised in the renal tubules higher levels are seen in patients with renal dysfunction.


C-Reactive Protein  

(Serum: 0 – 10 mg/l)‡ 
(Up to 4 days) 

Viral infection/AI disease:
11 – 49mg/l

Bacterial infection:
50 – 100mg /l

Major bacterial infection:
>100mg/l 

As CRP has a short serum half-life this acute phase protein is useful in distinguishing bacterial infections, inflammatory conditions, activity of rheumatoid arthritis and monitoring response to therapy. CRP may not be raised if a patient is on biological treatments such as anti-TNF.


Complement C3 and C4

(Serum: C3: 0.75 – 1.75 g/l)*
C4: 0.14 – 0.54 g/l)

(Up to 4 days) 

Measurement of C3 and C4 is of value in monitoring activity of SLE and in immune complex disease. C4 is of particular value in SLE and angioedema when levels are well below normal.


C1 (esterase) Inhibitor 

Immunochemical levels: 

(Fresh serum: 0.18 – 0.30 g/l)
(Up to 10 days) 

Functional activity 

(Fresh plasma (citrate): 70 – 130%)*
(Up to 28 days)

Please see page 6 in the CIS Handbook for collection procedure. 

Hereditary Angioedema

Autosomal dominant. Most cases have reduced serum C1Inh levels. In 10% of cases there are normal or elevated levels of C1Inh but this is functionally inactive. In hereditary angioedema C4 levels are almost always reduced and C1q levels are normal.

Acquired angioedema

Have reduced C1Inh levels and usually reduced levels of both C4 and C1q. Associated with B cell neoplasia.


Cryoglobulins

(Serum: Negative)
(Up to 10 days)

Positives may be typed:
monoclonal, polyclonal or mixed (IgG/A/M).

When cryoglobulins are associated with myeloma, Waldenströms macroglobulinaemia, or lymphoma they consist of one immunoglobulin isotype but may be mixed or polyclonal in other diseases such as connective tissue diseases. Patients with renal disease and a low C4 level or patients with unexplained cutaneous vasculitis should be screened for presence of circulating cryoglobulin. Please refer to special conditions of collection and despatch to laboratory (page 6 in the CIS Handbook).


Specific microbial antibodies (Functional antibodies) 

(Serum)
(Up to 28 days)

Pneumococcal ab protective level is 0.35 ug/ml for each serotype. 7/12 serotypes tested (4, 6B, 9V, 14, 18C, 19F, 23F) are present in both pneumovax II and Prevnar whilst a further 5 are present only in pneumovax II (1, 3, 5, 7f, 19a). A normal adult response to Pneumovax II is >0.35 ug/ml in 8/12 serotypes (6/12 in children aged 2 to 5 years).

Meningococcal C antibody protective level is 2.0 ug/ml. 

Hib ab protective levels - 1.0 ug/ml (long-term) and 0.15ug/ml (short-term). 

Tetanus antibody protective levels are 0.1 IU/ml (long-term) and 0.01 IU/ml (short-term).† 

Specific antibody testing can be helpful in patients with recurrent infections.

Specific antibody responses can be abnormal even if immunoglobulins are normal.

Antibody responses are normally assessed 4 – 6 weeks after vaccination.


Fungal antigens – Specific IgG antibodies 

(Serum: species specific ranges)*
(Up to 5 days)

Specific IgG antibodies directed against candida albicans, aspergillus fumigatus and micropolyspora faeni are available. Note: most adult women will have low levels of candida antibodies. IgG antibodies indicate previous exposure.


High sensitivity (ultrasensitive) C-Reactive Protein 

(Serum:)
Normal range: 0-5 mg/L*
(Up to 4 days)

Risk assessment guidelines:

hsCRP <1.0 mg/L = Low risk.
hsCRP 1.0-3.0 mg/L = Average risk.
hsCRP >3.0 mg/L = High risk. 

CRP measurement by high sensitivity methods can indicate the risk for future cardiovascular and peripheral vascular disease. Elevated values may be indicative of the prognosis of individuals with acute coronary syndromes or stable coronary disease. High sensitivity CRP (hsCRP) measurement should not be used as a substitute for assessment of traditional cardiovascular risk factors. Individuals with evidence of active infection, inflammation or trauma should not be tested for cardiovascular disease risk assessment by hsCRP measurement until these conditions have abated.


Immunoglobulins (IgG/A/M)  

(Serum: IgG:  6.00 – 16.00 g/l †
(adult):  IgA:   0.80 – 4.00 g/l †
IgM:  0.50 – 2.00 g/l) †
(Up to 4 days)

N.B. Reference ranges are age specific and may differ between ethnic groups 

Immunoglobulins are an essential request in recurrent infections, lymphoproliferative diseases including myeloma and all cases of ‘failure to thrive’. IgA deficiency occurs in 1:500 individuals but, transfusion reactions apart, may not be associated with disease. Polyclonal increases of IgG occur in chronic infection and inflammation, chronic liver disease and connective tissue diseases. Raised levels of IgM are found in acute inflammation and in primary biliary cirrhosis. (Markedly elevated IgM in the presence of mitochondrial antibodies is virtually diagnostic of PBC) Low levels of IgG and IgA may be due to loss (protein losing enteropathy or nephrotic syndrome), reduced synthesis (e.g. lymphoproliferative disorders or primary immunodeficiency) or excessive catabolism. Low immunoglobulins always require further investigation. Where appropriate details are supplied age and sex related normal levels are printed on the report. See also IgG sub-classes and functional antibodies.


IgG Subclasses  

(Serum: IgG1: 3.2 – 10.2 g/l †
adult: IgG2: 1.2 – 6.6 g/l †
IgG3: 0.2 – 1.9 g/l †
IgG4: 0.0 – 1.3 g/l)
(Up to 7 days) 

IgG subclass deficiency is mainly related to IgG1 and IgG2 where individuals may suffer recurrent infections.


Immunoglobulin D (IgD) 

(Serum:  0.05-0.20 g/L)
(Up to 14 days) 

Serum IgD is measured in the case of IgD myeloma and some forms of periodic fever syndrome.


Immunoglobulin E (IgE)

(Serum:  0 – 90 IU/ml, adult)
(Up to 7 days)

N.B. Reference ranges may differ between ethnic groups 

Serum IgE may be helpful in diagnosing atopic diseases however the reference range is very wide and levels do not correlate well with symptoms. Very high levels of IgE are seen both in atopic eczema and in parasitic infestations (especially S Mansoni) and may result in false positive specific IgE to a single allergen.


Lymphocyte cell markers

(Up to 4 days) 

A wide range of lymphocyte markers for assessment of immunodeficiency and lymphoproliferative diseases are available. A separate request form is in use for cell markers.


Lymphocyte function tests

(Up to 28 days)

Special sample requirements. Please discuss with Consultant Immunologist.


Mast cell tryptase 

(Serum: 0 – 13.5 mg/l)

Samples may be kept at room temperature for shipping purposes for 2 days.

(Up to 5 days – unless agreed as urgent by telephone) 

For suspected cases of anaphylaxis. A clotted blood sample should be taken immediately, four hours and at 24 hrs post reaction. It is often difficult to give an immediate sample if patient is not in hospital. Tryptase release in IgE mediated allergy peaks at 4 hours and then decline. Please indicate time/date of adverse reaction and time/date of samples.

All patients who have suffered anaphylaxis should be referred to a specialist allergy clinic (NICE guideline http://www.nice.org.uk/guidance/cg134).

A persistently raised tryptase may indicate mastocytosis or a mast cell activation disorder.


Neutrophil respiratory burst test

(4 days)

This is a diagnostic test for chronic granulomatous disease. Please arrange with laboratory


Paraprotein (Monoclonal protein, M-protein) quantitation

Reported in g/l.
(Up to 4 days)

Levels of monoclonal IgG, IgA, IgM, IgD and in some instances IgE are measured immunochemically. Immunofixation of presentation sample defines both the isotype and light chain type. Follow up specimens will be subjected only to electrophoresis unless immunofixation is required to confirm complete response. (See ‘guide to appropriate use of tests at the front of this booklet). The presence of an M-protein (paraprotein) should prompt investigation of B cell malignancy, particularly myeloma, (IgG, IgA) and lymphoplasmacytoid lymphoma (IgM). Monoclonal gammopathy of uncertain significance (MGUS) is found in one or more percent of the general population over the age of 50 years.


Serum specific IgE (allergen specific IgE)

(Serum: 0 – 0.35 kU/l)*
(Up to 7 days, if in stock)

N.B. Total IgE will be carried out on all Serum specific IgE requests unless an IgE level is stated at the time of request.

Assays for the detection of circulating IgE antibodies directed against specific antigens are available to a wide range of allergens. Tests for common substrates include animal fur or dander, house dust mite, tree and grass pollens, moulds, feathers and an extensive range of food substances including a variety of nuts and are performed in house. Requests for more unusual allergens are sent to Sheffield.


Serum electrophoresis

(Serum: Clinical comment will accompany each report)

(Up to 4 days)

Sera are screened for qualitative abnormalities in proteins especially of the immunoglobulins. Scans demonstrating a monoclonal band are automatically followed up using immunofixation to determine both the isotype and the light chain of the monoclonal protein. Other typical patterns seen on electrophoresis may indicate evidence of acute phase responses, immunodeficiency, etc.

Where myeloma is suspected urine and serum should be sent together.


Serum immunoglobulin free light chains (FLC)

Kappa 3.30 – 19.40 mg/l *
Lambda 5.71 – 26.30 mg/l *
Kappa / Lambda ratio 0.26 – 1.65 *
(Up to 4 days)

This assay may be inaccurate at levels <0.9mg/l. In a small proportion of patients with high serum FLC levels, false negative results may occur as a result of “antigen excess”. Any anomaly between the serum FLC results and other laboratory tests and/or clinical evidence should be reported to the laboratory for re-testing the serum FLC.

Normal plasma cells make more immunoglobulin light than heavy chains and secrete free light chains in amounts detectable in serum (estimated to be 0.5g/day). Serum free light chains are removed by glomerular filtration with a half-life of a few hours. They are not easily detectable in urine until the threshold for tubular reabsorption is exceeded (10 – 20g / day).

Serum FLC measurements are recommended in assessment of all plasma cell dyscrasias and in B cell lymphoproliferative diseases. They are particularly important in diagnosis and management of light chain only myeloma.


Key for reference ranges

Where a quantitative reference range is provided the origin of that range is as follows:

*

Manufacturers reference range (verified in-house)

National (or international) reference laboratory range (verified in-house)

Internally generated reference range

§

Send away test with accompanying (referral laboratory derived) range

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Routine and Urgent Assay Processing

Daily assays

As a general guide, most immunochemistry, indirect immunofluorescence, electrophoresis, immunophenotyping and some neuroimmunology assays are carried out on a daily basis.

Batched assays

Non urgent, expensive or labour intensive assays which are batched on a less frequent basis include:

Cardiolipin/B2GP1 antibodies

Wednesday

dsDNA antibody ELISA

2 or 3 times a week

ENA antibodies

Thursday

Intrinsic factor antibodies

Monday

Functional C1Inh

Once a week (see page 6 in the CIS Handbook for collection requirements)

Organ specific antibodies

Friday

IgG subclasses

Twice a week

MPO/PR3 antibodies

2 or 3 times a week

GAD antibodies

Every 2 weeks

M2 mitochondrial ELISA

Once a week

Urgent assays

(e.g. Tau protein, MPO/PR3/GBM abs, acute leukaemia immunophenotyping)

Some assays are available with a reduced turnaround time on discussion with a member of the senior laboratory staff. Urgent specimens must be discussed with the laboratory and handed over to a member of staff in specimen reception within the CIS at the medical school. The request form must be clearly marked “Urgent” and with which member of staff the request was discussed. The sample must arrive before 2pm. Contact details (ideally a mobile phone number) for the requesting clinician must also be supplied to enable results to be communicated urgently.

Prior warning for urgent requests is essential. It should be noted that urgent requests may incur an additional charge and where an initial qualitative result is provided these will be followed up with quantitative assays when the next routine batch is processed.

General Specimen Collection Requirements

When sending specimens to the laboratory the following should be noted:

  • Different samples require different blood tubes. For example; red topped tubes (in the Vacutainer system used at UHB) allow blood to clot for tests that require serum.
  • Some complement components are unstable. For functional complement assays (CH50/AP50) send 5ml of blood in a red topped tube (for C3d use EDTA blood) immediately after venepuncture. For Functional C1Ihn please send 5ml blood in a sodium citrate (blue top) tube. For distant clinics the serum or plasma should be separated within the hour, frozen and sent to the laboratory to arrive frozen.
  • For cryoglobulins blood MUST be taken into a warm red top Vacutainer tube and maintained at no less than 37°C (and up to 40°C). For UHB samples, the laboratory will bring a pre-warmed container with electronic temperature tracking probe and phlebotomy equipment and provide advice on sampling – please contact lab prior to performing this test to arrange.
  • T cell antigen receptor & immunoglobulin gene rearrangement studies: Please supply blood or bone marrow samples drawn into an EDTA bottle (Please note: heparinised material may interfere with PCR and will NOT be processed).
  • T-SPOT TB: Two 6ml Lithium Heparin specimens are required for this assay. Use Cell Markers request form. Samples must arrive in the laboratory before 2pm on the same day as they are drawn. (Tuesdays only: On other days this assay may be available under special circumstances and by prior arrangement only). This assay must be booked with laboratory staff before sending.
  • All samples MUST be shipped to the department in appropriate packaging. For transport via road, rail and/or air regulations must be followed and UN3373 compliant packaging used.

For all immunophenotyping and cell function assays please see the notes towards the end of this CIS handbook under ‘Immunophenotyping’ (p29) and ‘Cell function assays’ (p33).

All high-risk specimens and their accompanying forms MUST be clearly labelled. Samples are not tested on site if they are from a patient with suspected CJD or vCJD. Where such samples are forwarded to an alternative laboratory, turnaround times will be longer.

Please also note special requirements for cell work and neuroimmunology requests both of which have separate request forms.

A Brief Guide to Common Immunology Tests

Anti-nuclear antibodies (ANA)

This test is most useful in situations where SLE is suspected – as ANA negative SLE is rare. ANA is also associated with a number of connective tissue diseases and therefore high titre ANA results may be considered supportive evidence of this. However, positive ANA (particularly at low titre) may also be seen after infection or even in asymptomatic individuals (especially older people and females).

SLE in pregnancy or with planned pregnancy

These patients should have anti-cardiolipin (ACL) antibodies tested and their Ro antibody status determined. Ro antibodies are specific for a type of extractable nuclear antigen (ENA), and are relevant in pregnancy because they are associated with congenital heart block.

Anti-neutrophil cytoplasmic antibodies (ANCA)

This test is most useful in situations where small vessel vasculitis is suspected. ANCA have different specificities, of which the most clinically relevant are MPO and PR3 antibodies. These are particularly associated with microscopic polyangiitis, eosinophilic granulomatosis with polyangiitis (EGPA) and granulomatosis with polyangiitis (GPA). However, clinicians should be mindful that vasculitis is not excluded by a negative ANCA result.

Allergy tests

Specific IgE tests to individual allergens can support a diagnosis of allergy in a patient who has compatible symptoms. However, specific IgE antibodies are not useful as a screening test because they can be positive in asymptomatic patients. These patients may be sensitized but not necessarily allergic. These tests only have a role in type I hypersensitivity reactions, which are characterised by a rapid onset of symptoms following exposure.

Suspected Myeloma

Immunoglobulins should be measured (this includes levels of IgG, IgA and IgM). Electrophoresis is performed to identify whether there is a paraprotein. If present then the type and size of the paraprotein is determined by immunofixation and densitometry respectively. Not all myeloma patients will secrete an intact paraprotein and therefore kappa and lambda serum free light chains are also typically requested

How do I order a test?

Labatory testing and results

For general telephone enquiries and results: +44(0)121 414 4069
Fax: +44(0)121 414 3069

Non NHS Organisations e.g. COVID-19 Antibody testing (form attached): Clin.imm@nhs.net

Non NHS individuals please arrange through your General Practitioner.

Laboratory Manager: +44(0)121 414 3092

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