The sample requirements described below refer to the analytical methods currently used by Laboklin for the measurement of various analytes. As analytical methods continue to evolve, these requirements may change over time. If in doubt, contact our laboratory before sample collection.
As a general rule, marked haemolysis and lipaemia should be avoided. Common causes of haemolysis include prolonged venous occlusion, difficult venepuncture, clotting in the refrigerator, and failure to separate serum or plasma before shipment. Fasting blood samples are generally recommended for hormone measurements to minimise lipaemia.

 

Thyroid Gland

T4 (Total T4, TT4; Free T4, fT4) and T3

Indications: Screening for hypothyroidism; screening and diagnosis of hyperthyroidism.

Important considerations before sample collection: T4 concentrations may be reduced by non-thyroidal illness (NTI) and by various medications. Free T4 is somewhat less affected by these factors, but its concentration may also be decreased by NTI and certain drugs. Differentiation from genuine hypothyroidism requires further diagnostic investigation.

Analytical methods: Measurement can be performed using two different methods.

1. CLIA (Chemiluminescent Immunoassay): This assay is based on the immunological binding of the T4 molecule. For initial screening, this more cost-effective method is often sufficient.
2. LC-MS/MS (Liquid Chromatography-Tandem Mass Spectrometry): This method is increasingly regarded as the gold standard. It is considered particularly precise because it measures only the T4 molecule itself, thereby avoiding interference from metabolites or antibodies. At Laboklin, LC-MS/MS testing is available for T4, T3 and rT3 (see below).

Sample material: Serum (mandatory for LC-MS/MS; preferred for CLIA).

Good to Know: TT4 and fT4 remain stable at room temperature for three to five days. For fT4 it is being discussed that dissociation of T4 from its binding proteins may occur after more than five days and/or when exposed to high ambient temperatures. This may result in elevated fT4 concentrations, potentially leading to misinterpretation of results.

TSH

Indications: Investigation of hypothyroidism; treatment monitoring.

Important considerations before sample collection: In the presence of a low T4 concentration and compatible clinical signs, an elevated TSH concentration generally confirms hypothyroidism. However, approximately 30% of hypothyroid dogs have low TSH concentrations. In such cases, further diagnostic investigations are required to differentiate between NTI and hypothyroidism.

Analytical method: CLIA

Sample material: Serum

Good to Know: TSH is stable for 24 hours at room temperature and for seven days when refrigerated (2 to 8°C).

Reverse T3 (rT3)

Indications: Aid in differentiating between NTI and hypothyroidism in dogs with reduced T4 concentrations and no increase in TSH; support in cases of suspected hyperthyroidism in cats when T4 concentrations are not elevated as expected.

Important considerations before sample collection: rT3 is a highly promising parameter. However, interpretation should always be performed in the clinical context and in combination with other thyroid parameters.

Analytical method: LC-MS/MS

Sample material: Serum

Good to Know: rT3 remains stable at room temperature for up to 14 days.

Autoantibodies (against Thyroglobulin, T3 and T4)

Indications: Suspected interference with T4 measurement by CLIA; breeding health screening and breeding approval programmes.

Important considerations before sample collection: Immunosuppression (medication) and immune stimulation (for example vaccination) may alter antibody concentrations. This should be taken into account when interpreting results.

Analytical method: ELISA. Results are reported either quantitatively (percentage proportion in the sample) or qualitatively (positive/negative).

Sample material: Serum

Good to Know: Antibodies are relatively stable. However, samples should be transported refrigerated if transport times exceed 48 hours. Sample quality is particularly important for antibody testing. Serum must be cleanly separated, and haemolysis and lipaemia must be avoided.

 

Parathyroid Gland

Parathyroid Hormone (PTH)

Indications: Investigation of hyperparathyroidism or hypoparathyroidism.

Important considerations before sample collection: As interpretation depends on correlation with the blood calcium concentration at the time of sampling, PTH and calcium should be measured from the same serum sample or from serum collected at the same sampling time.

Sample material: Serum

Analytical method: CLIA

Good to Know: PTH is unstable. Consequently, proper pre-analytical handling is of great importance. The sample should be allowed to clot for no longer than 30 minutes, then centrifuged immediately and the serum transferred into a separate tube. The separated serum must be stored and transported frozen. At a minimum, shipment should be carried out using pre-frozen cooling packs; transport on dry ice is ideal.

 

Adrenal Gland

Cortisol

Indications: Screening parameter for ruling out hypoadrenocorticism; measurement as part of the ACTH stimulation test and the dexamethasone suppression test.

Important considerations before sample collection: Cortisol secretion increases in response to both emotional and physical stress. This can affect the results of functional tests. In particular, the dexamethasone suppression test should be performed only after concurrent diseases have been resolved and under conditions that are as stress-free as possible.

Analytical methods: CLIA; LC-MS/MS may be used for specific clinical questions. In routine diagnostics, cortisol is generally measured using immunological assay methods (CLIA). In human medicine, however, LC-MS/MS is increasingly becoming established as a highly specific and precise method for cortisol measurement.

Sample material: Measurement can be performed using plasma. However, serum is preferred to ensure standardisation of hormone testing. In particular, the same sample material should always be used for follow-up measurements and functional tests to ensure consistent and comparable results.

Good to Know: Cortisol in serum samples is generally stable at room temperature for 3 to 5 days. Marked lipaemia may affect cortisol measurements performed using immunoassays. Although fasting is not strictly required for functional testing, it may be advisable to feed only a small and/or low-fat meal before testing in order to minimise lipaemia-related interference.

Urine Cortisol-to-Creatinine Ratio (UCCR)

Indication: Screening for possible Cushing’s syndrome.

Important considerations before sample collection: Urine samples for UCCR determination should ideally be collected in the morning and in a stress-free environment, with at least three (better five) days between sample collection and a veterinary visit. Before submission, urinary tract infection should be excluded by performing urinalysis, including sediment examination.

Analytical method: For one of the immunoassays widely used in veterinary medicine for urinary cortisol measurement (CLIA, Immulite 2000®, Siemens, Germany), changes made by the manufacturer to the assay substrate have led to concerns regarding the reliability of results obtained from canine urine samples. The assay used by Laboklin is not affected by this issue.

Endogenous ACTH (eACTH)

Indications: Further differentiation of Cushing’s syndrome and hypoadrenocorticism (Addison’s disease).

Analytical method: CLIA

Sample material: EDTA plasma

Good to Know: Endogenous ACTH is unstable in biological samples, particularly in dogs. It is rapidly degraded by endogenous enzymes in the blood, which may lead to falsely low results. When stored frozen at -20°C in separated plasma, eACTH remains stable for extended periods. Blood should therefore be collected into pre cooled EDTA tubes, ideally chilled in a freezer at -20°C before sampling. Centrifuge and separate immediately, transfer plasma to an uncoated tube and ship frozen. Ideally, the sample should arrive at the laboratory still frozen. Glass tubes are not suitable, as eACTH binds to glass surfaces, which can reduce the measured concentration.

Aldosterone

Indication: Investigation of hyperaldosteronism.

Important considerations before sample collection: The patient should be adequately hydrated, as the renin-angiotensin-aldosterone system (RAAS) is influenced by hypovolaemia and dehydration.

In azotaemic patients, measurement of aldosterone alone is not sufficient for the diagnosis of primary hyperaldosteronism, as aldosterone concentrations may also be elevated in renal disease. Feeding time can have an effect on the RAAS; therefore, collection of a fasting blood sample is recommended.

Sample material: Serum

Analytical method: LC-MS/MS. This method is considered highly precise and superior to immunological assay methods. It is important to note that LC-MS/MS typically measures lower aldosterone concentrations than other analytical methods. This has been taken into account in the reference intervals provided by Laboklin.

Good to Know: The sample should be as fresh as possible and no older than 48 hours. Allow to clot for ≤30 minutes, centrifuge immediately and transfer serum to a separate tube. The separated sample should be cooled as quickly as possible and must arrive at the laboratory under refrigerated conditions.

RAAS Profile (Calculation of Renin Activity, ACE Activity and the AA2 Ratio) 

Indication: Investigation of hyperaldosteronism in patients with concurrent azotaemia.

Important considerations before sample collection: The patient should be adequately hydrated, as the renin-angiotensin-aldosterone system (RAAS) is influenced by hypovolaemia and dehydration. Feeding time can have an effect on the RAAS; therefore, collection of a fasting blood sample is recommended.

Sample material: Serum

Analytical method: Renin activity is calculated following measurement of aldosterone, angiotensin I and angiotensin II by LC-MS/MS. The method has been validated for both cats and dogs.

Good to Know: Angiotensin I and angiotensin II are unstable and must not be exposed to temperatures of 4 to 8°C. Consequently, the sample must arrive at the laboratory frozen. The sample should be allowed to clot for no longer than 30 minutes, then centrifuged immediately and the serum transferred into a separate tube. The serum needs to be frozen immediately at -20°C. The sample must not be stored in a refrigerator prior to freezing and, once frozen, must not be allowed to thaw before arrival at the laboratory (!). Shipment on dry ice is mandatory. 

Catecholamines (Metanephrine, Normetanephrine)

Indications: Investigation of a pheochromocytoma or catecholamine-producing paraganglioma.

Important considerations before sample collection: Catecholamines are released in response to stress. Consequently, stressful situations may lead to increased catecholamine secretion. Urinary measurement is less affected by episodic catecholamine release than blood measurement. Urine samples may be collected either by cystocentesis or as a free-catch sample. Collection in the home environment is not mandatory. However, a urinary tract infection should be excluded before sample submission by performing a urinalysis, including sediment examination.

Medications such as phenoxybenzamine, metoclopramide, β-blockers, calcium channel blockers or sympathomimetics (epinephrine,
dopamine, dobutamine, phenylpropanolamine and terbutaline) may cause falsely elevated results. Particular attention should be paid to ensuring that sample collection takes place before initiation of any potential treatment with phenoxybenzamine.

Analytical method: LC-MS/MS

Sample material: EDTA plasma or urine (free-catch or cystocentesis sample)

Good to Know: Catecholamines are unstable in blood samples. Blood should therefore be centrifuged immediately and the plasma separated without delay. After transfer into an uncoated sample tube, the plasma should either be stored refrigerated at 4°C, where it remains stable for up to three days, or frozen at -20°C. Samples should be shipped under refrigerated conditions and must arrive at the laboratory chilled. Shipment with pre-frozen cooling packs is the minimum requirement; transport on dry ice is ideal. In urine, catecholamines remain stable for several days even at room temperature. The acidification of urine samples that was previously recommended is no longer performed. Studies in both human and veterinary medicine have demonstrated that acidification is not necessary.

 

Pituitary Gland

Insulin-Like Growth Factor 1 (IGF-1)

Indications: Acromegaly (hypersomatotropism) and pituitary dwarfism (hyposomatotropism).

Analytical method: CLIA

Sample material: Serum

Good to Know: The sample should be allowed to clot for no longer than 30 minutes, then centrifuged immediately and the serum transferred into a separate tube. Provided that the blood sample is centrifuged and separated promptly, IGF-1 remains stable for 24 hours at room temperature and for up to seven days at 4°C. Cooling of the sample and shippment under refrigerated conditions is recommended, especially if prolonged transportation times are expected.

 

Endocrine Pancreas

Insulin

Indication: Insulinoma

Important considerations before sample collection: Blood sampling should be performed during documented hypoglycaemia (blood glucose < 3.3 mmol/L or < 60 mg/dL). Glucose and insulin measurements must be performed on serum obtained from blood collected at the same time.
Ideally, collect blood from a large vessel (e.g. the jugular vein) using a large-bore needle to minimise haemolysis and facilitate rapid collection.

Analytical method: CLIA (dog), ELISA (cat)

Sample material: Serum

Good to Know: Insulin is unstable in serum at room temperature and may undergo significant degradation within a few hours. The blood sample should therefore be allowed to clot for no longer than 30 minutes, then centrifuged immediately and the serum transferred into a separate tube to minimise degradation. The sample should be stored refrigerated at 4°C. If analysis cannot be performed within 24 hours, freeze serum at -20°C (-80°C for long-term storage). Refrigerated shipment must be ensured using pre-frozen cooling packs; for longer transport times, shipment on dry ice is recommended. The sample should arrive at the laboratory refrigerated (4-8°C). As haemolysis may result in the release of insulin-degrading enzymes, it should be avoided whenever possible.

Jennifer von Luckner, Ruth Klein

 

Our Endocrinology Services Include

• Individual hormone assays
• Functional endocrine testing
• Specialised diagnostic profiles

In the same study, treatment success rates for chronic diarrhoea in dogs, cats, and horses ranged between 61–85%, and for chronic rhinitis in cats from 70% (8). In horses, autovaccines can also be used in cases of free faecal water (9).

In another study on idiopathic recurrent pyoderma in dogs, administration of a Staphylococcus pseudintermedius-based autovaccine in addition to antibiotic therapy resulted in significantly improved pyoderma scores compared with the antibiotic-only group (10). Success rates for this indication in further studies between from 77–88% (11–13).

Abscesses in rabbits caused by highly virulent S. aureus strains decreased in size within two weeks of autovaccine treatment, although they were not completely eliminated (14).

Unlike conventional vaccines, autovaccines do not contain adjuvants. The use of immunomodulators to enhance the cellular immune response prior to administration of autovaccines for staphylococcal infections has been described (6).

 

Mechanisms of action of autovaccines

Autovaccines do not act through direct elimination of the pathogen, but rather through a general stimulation of the immune system, enabling the host to eliminate the pathogen itself.

Sir Almroth Edward Wright already hypothesised an increase in phagocytic activity following autovaccine therapy (4, 5). Although the exact mechanism of action has not yet been fully clarified, it is assumed that the innate immune system is activated first, leading to a non-specific immune response and the recruitment of phagocytic defence cells such as macrophages to the site of infection.

This is followed by activation of the adaptive immune response, in which T and B cells are involved and both cellular and humoral defence mechanisms are triggered (6, 15). This distinguishes autovaccines from conventional vaccines, which primarily induce a humoral immune response (6).

Furthermore, it has been shown that autovaccine treatment increases the production of pro-inflammatory cytokines, thereby enhancing overall immune activity (16). Orally administered autovaccines, for example in cases of bacterial diarrhoea or other gastrointestinal infections, also lead to an increase in secretory antibodies (IgA), which protect the intestinal epithelium from bacterial adhesion and thereby strengthen the mucosal barrier (17, 18).

Interestingly, autovaccines may also influence the bacterial population targeted within the host. In one study, strains with a reduced genetic repertoire for survival in the host predominated after treatment (19). This may be advantageous for therapy.

 

Autovaccines at Laboklin

Autovaccines at Laboklin can be produced for companion animals that are not used for food production. Farm animals intended for food production are excluded.

The exact ordering process for an autovaccine consists of several steps, including the initial bacteriological examination, the required documentation, as well as manufacturing and dispatch (Fig. 1).

Autovaccines should not be administered to young animals under one year of age.

The routes of administration include injection vaccines for chronic skin or ear infections, inhalation vaccines for chronic respiratory infections, oral (swallow) vaccines for chronic diarrhoea, and combination vaccines with both oral and injectable components for infections of the urogenital tract.

An overview shows which autovaccine types are most commonly produced for the respective animal species depending on the clinical presentation (Fig. 2).

The administration period for the different autovaccines is usually 3–4 weeks. A prerequisite for production is completing the Laboklin prescription forms and, in Germany, the submission of the pharmacy authorisation for the veterinary practice’s in-house pharmacy.

An autovaccine can be produced when no commercially available vaccines exist against the pathogen. Regulation (EU) 2019/6 on veterinary medicinal products governs the manufacture, prescription, and supply of autovaccines at EU level (20). It also states that the attending veterinarian must ensure that the isolated pathogen is administered as an autovaccine only to animals belonging to the same epidemiological unit or, if at different locations, to those with a verified epidemiological link (21).

Once the legal requirements are fulfilled, the autovaccine is produced at Laboklin. To ensure quality and safety, this includes not only inactivation of the pathogens and adjustment of the microbial concentration, but also a two-week sterility control in accordance with the European Pharmacopoeia (22).

A maximum of four pathogens can be included in a single autovaccine. In general, autovaccines can be produced for aerobically growing bacteria, with the exception of aerobic spore-forming bacteria.
Obligate anaerobic bacteria, viruses, and fungi are also excluded.

Before manufacturing an autovaccine, it should be assessed whether the bacteria isolated in the microbiological examination are potentially pathogenic for the specific anatomical site. An autovaccine targeting commensal flora is not useful.

The six most common pathogens for each autovaccine type are summarised in an overview (Fig. 3). In cases of recurrence, a follow-up vaccine can be produced within one year using bacteria obtained from the microbiological examination. In some cases, a repeat bacteriological analysis is useful to determine the current spectrum of pathogens.

In particular, pyoderma cases have an expected recurrence rate of around 20% (7, 8).

An autovaccine can also be used as an adjunct to antibiotic therapy. However, concurrent antibiotic therapy is not recommended in the case of oral (swallow) vaccines. This is because antibiotic treatment may alter the gut microbiota and mucosal immune homeostasis, potentially reducing the effectiveness of the IgA antibody response induced by the autovaccine.

Adverse drug reactions associated with autovaccines are relatively rare. However, redness and swelling at the injection site may occur.
Systemic reactions such as fever, increased respiratory rate, and apathy can also be seen (6).

It should be noted that underlying diseases may influence the effectiveness of the autovaccine.

 

Conclusion

Autovaccines represent an important alternative or adjunct to antibiotic therapy in chronic diseases of companion animals, particularly in the context of increasing antimicrobial resistance.

 

Johannes Kupke, Martina Krapf

 

Services related to this topic:

• Bacteriology (aerobic)
• Oral vaccine
• Injectable vaccine
• Combination vaccine
• Inhalation vaccine

The Laboklin expert panel addresses a wide range of clinically relevant questions. On the topic of feline hyperthyroidism, four endocrinologists and one surgeon came together to share their expertise with us.

The following experts took part in the discussion: Prof. Dr. Felicitas Boretti, Dipl. ACVIM and ECVIM-CA, Head of Endocrinology, Clinic for Small Animal Medicine, Vetsuisse Faculty, University of Zurich; Dr. Katarina Hazuchova, PhD, Dipl. ECVIM-CA, Senior Physician in Internal Medicine, Small Animal Clinic, Giessen; Prof. Dr. Mirja Nolff, Dipl. ECVS, Head of Soft Tissue and Oncological Surgery, Clinic for Small Animal Surgery, Vetsuisse Faculty, University of Zurich; PD Dr. Astrid Wehner, Dipl. ECVIM-CA, Senior Physician in Internal Medicine with a focus on Endocrinology, LMU Munich; PD Dr. Florian Zeugswetter, Head of the Endocrinology Department, University Clinic for Small Animals, Vienna.

Before moving on to diagnostics and therapy, we briefly review the development of feline hyperthyroidism: PD Dr. Zeugswetter explains that the normal feline thyroid gland contains a subpopulation of thyrocytes with a high growth potential. When these are stimulated by certain external factors, hyperplasia occurs. The hyperplastic cells eventually begin to proliferate autonomously, and adenomatous hyperplasia progresses to adenomas and, in rare cases, adenocarcinomas.

The exact triggers of adenomatous hyperplasia are not known. The sharp increase in cases since the disease was first described in 1977 suggests the involvement of environmental factors. Factors as-sociated with the condition in numerous studies in-clude indoor-only housing, canned food, fish-based diets, large fluctuations in dietary iodine, cat litter, and flavonoids (e.g. in soya). Polyphenols (BPA) and polybrominated diphenyl ethers (PBDEs) in food, drinking water, or the environment are also thought to play a role.

PD Dr. Zeugswetter then reviews the most important clinical signs of feline hyperthyroidism.

The main symptoms he mentions are weight loss despite a good appetite, polyuria/polydipsia, scruffy coat, marked restlessness, and increased vocalisation. Gastrointestinal signs such as vomiting and diarrhoea are also common; sometimes diarrhoea may be the only symptom.

This immediately raises the question: which less typical signs should not be overlooked?

Dr. Katarina Hazuchova points out that cats with hyperthyroidism may show pronounced weakness with cervical ventroflexion. Hyperthyroidism should also be considered as a differential diagnosis in cases of polypnoea, as it may reflect increased oxygen demand. Likewise, thyroid function should always be checked in cats with newly developed heart murmurs or other signs of cardiac disease.

The same applies to cats with epileptiform seizures or sudden blindness.

PD Dr. Astrid Wehner emphasises that cardiac changes induced by hyperthyroidism are often not initially clinically relevant and may normalise again once the hyperthyroidism has been successfully treated. Cardiac biomarkers such as (NT-)proBNP or troponin I can be helpful for assessment. If these values do not decrease later on despite well-controlled hyperthyroidism, a cardiological examination should be carried out in order to further classify any underlying heart disease and treat it if necessary.

The laboratory changes that may indicate the presence of hyperthyroidism are discussed by Prof. Dr. Felicitas Boretti. She explains that in cats with elevated liver values and compatible clinical signs, hyperthyroidism should be included in the differential diagnoses. Both ALP (alkaline phosphatase) and ALT may be increased; in practice, ALT is more frequently elevated, although it remains non-specific, whereas an increase in ALP is more typical in the context of hyperthyroidism.

Elevated ALP and phosphate levels in combination with normal renal parameters can be explained by increased activity of bone-specific isoenzymes, reflecting increased bone turnover, and should prompt targeted thyroid diagnostics when clinical findings are compatible.

In most cases, the diagnosis is made based on an increased serum T4 concentration. PD Dr. Florian Zeugswetter addresses the question of how the cat’s age influences the reference interval. In kittens, T4 concentrations are physiologically very high and gradually decrease with age. For cats between approximately 1 and 9 years of age, standard reference ranges are generally appropriate.

In older cats, the reference intervals may, however, be set somewhat lower. Some laboratories have taken this into account by using a lower upper reference limit, as hyperthyroidism is primarily a disease of older cats. When interpreting T4 results, both the cat’s age and the upper reference limit of the respective laboratory should be considered.

This raises the question of how often cats with hyperthyroidism still show a T4 concentration within the reference range. Prof. Dr. Felicitas Boretti points out that this can occur, particularly in the early stages of the disease. If T4 is only mildly increased, natural fluctuations may temporarily bring it back into the reference range, although it is usually in the upper part of the range. Non-thyroidal illness can also lower T4 to the extent that it temporarily falls back within the reference interval.

In cases of unclear findings, PD Dr. Florian Zeugs-wetter recommends repeating the T4 measurement after 2–4 weeks. If there is a strong clinical suspicion of hyperthyroidism but T4 is within the upper reference range, measurement of free T4, endogenous TSH, and reverse T3 can be helpful.

However, the limitations of these parameters must be considered: fT4 may be increased in non-thyroidal illness (NTI), making it relatively non-specific; a TSH concentration within or above the reference range makes hyperthyroidism highly unlikely, but a non-detectable TSH is also common in healthy cats; reverse T3, an inactive metabolite of T4, reliably increases in hyperthyroidism, although further studies are still needed.

If these advanced parameters also fail to clarify the situation, the next step in the diagnostic work-up is the T3 suppression test, or preferably thyroid scintigraphy.

The question is raised as to when a thyroid carcinoma should be suspected instead of benign adenomatous hyperplasia or an adenoma. PD Dr. Astrid Wehner names rapidly growing, solid masses (if not cystic), as well as cats that are difficult to stabilise and require unusually high doses of antithyroid medication, as warning signs. Dr. Katarina Hazuchova adds that these are often patients that have been hyperthyroid for a longer period of time. PD Dr. Florian Zeugswetter also describes palpation findings as indicative: benign nodules tend to be soft and easily movable, whereas firm, poorly movable and larger changes are more suggestive of carcinoma. Scintigraphy is unfortunately less reliable for definitive differentiation than one might expect.

In terms of therapeutic options, the question of prognosis is raised. PD Dr. Astrid Wehner reports encouraging outcomes here. Overall, the prognosis is favourable, although it also depends on the cat’s age and, of course, any comorbidities.

Curative treatment options such as radioiodine therapy are associated with significantly longer life expectancy, lower recurrence rates, and fewer treatment-related side effects. After radioiodine therapy, the median survival time is

approximately 3.7 to 4.0 years. In comparison, the prognosis under medical treatment with antithyroid drugs is less favourable. In a study involving 47 cats treated long-term with thiamazole/methimazole, the median survival time was 2.0 years.

This difference can be explained by several factors. Owners may, over time, no longer be able to consistently administer medication to their cat, or the hyperthyroidism may no longer be adequately controlled medically. With prolonged treatment using antithyroid drugs, resistance may also develop. In addition, adenomatous changes may, over time, progress to carcinoma.

Prof. Dr. Mirja Nolff lists the main indications for thyroidectomy as cats that are difficult to stabilise, in which medication administration is not feasible, those experiencing adverse effects from antithyroid drugs, cats with large nodules, or cases where carcinoma is suspected.

It becomes critical when bilateral surgery is required and the parathyroid glands cannot be preserved.

Reimplantation is not reliably successful, so postoperative hypoparathyroidism must be anticipated. Postoperatively, ionised calcium should be monitored very closely. Another serious but rare complication is laryngeal paralysis.

Prof. Dr. Nolff emphasises that, compared with radioiodine therapy, unilateral thyroidectomy performs less well in achieving euthyroidism. This is because both thyroid lobes are often affected. She therefore recommends informing cat owners that unilateral surgery may not fully resolve the clinical signs. In such cases, removal of the second thyroid lobe may still be necessary if radioiodine therapy is not available. Any resulting hypothyroidism is usually manageable.

The question of whether dietary therapy is an option is addressed by PD Dr. Astrid Wehner. A marked reduction in iodine intake can suppress thyroid hormone production sufficiently to achieve euthyroidism. A specific prescription diet is available. Simply avoiding foods with high iodine

content (fish, seafood) is not sufficient. The time required to achieve euthyroidism varies considerably (28 to 180 days). The diet must be strictly, completely, and permanently adhered to. Unfortunately, this is often the reason for long-term treatment failure. Overall, T4 concentrations may fluctuate and, in some cases, remain elevated.

Antithyroid drugs remain the most commonly used treatment option. Approved veterinary products include carbimazole as a sustained-release formulation and thiamazole (methimazole).

Sustained-release carbimazole can be given once daily according to the manufacturer, while thiamazole preparations are usually administered twice-daily.

Prof. Dr. Felicitas Boretti explains that a trial of once-daily dosing is generally possible. However, in cats with very high initial T4 levels, twice-daily

administration usually leads to faster stabilisation. If T4 levels fall too low at a low total dose, switching to once-daily administration may be appropriate.

Another option is a veterinary-licensed, individually dosed syrup formulation.

Regarding transdermal ointment treatment, PD Dr. Astrid Wehner and PD Dr. Florian Zeugswetter are rather cautious. This formulation has several disadvantages: the active substance, which is teratogenic to unborn children, may come into contact with the owner more easily; absorption is not always consistent; and residual cream in the ear may further affect uptake. In the future, a nanoparticle carrier ointment may become available in Germany, and studies on this are already underway.

PD Dr. Wehner also addresses hyperthyroidism and kidney function. In hyperthyroid cats, there is an increased circulating blood volume and consequently increased renal perfusion. This leads to glomerular hypertension, possibly accompanied by systemic hypertension, which can damage the kidneys and/or contribute to the progression of pre-existing renal disease, as proteinuria may also result. At the same time, glomerular filtration rate increases due to the higher intraglomerular pressure, resulting in lower blood concentrations of renal filtration markers such as creatinine and SDMA.

It is possible that a renal disease only becomes evident after normalisation of thyroid function. The experts agree that in cats that are already azotaemic at the time of diagnosis, treatment with antithyroid drugs should initially be started at a reduced dose. As a rule of thumb, this is about half of the standard dose, or slightly less depending on the severity of the azotaemia, in order to assess the kidneys’ response to therapy.

Radioiodine therapy (RIT) is considered the optimal treatment option. Dr. Katarina Hazuchova explains what needs to be considered when referring a patient. The cost of standard treatment is around €2000 net, and cats usually remain hospitalised for 5–7 days. A cardiological assessment should be performed beforehand; in some centres, full diagnostic work-up including abdominal ultrasound is also required. Antithyroid medication should be discontinued 7 days before treatment, and an iodine-restricted diet 14 days prior. After RIT, cats must remain indoors for four weeks; if pregnant women, breastfeeding mothers, or babies/young children live in the household, they must not come into contact with the cat. Cats that may require intensive care or do not maintain adequate food intake are not suitable candidates for this treatment.

Regarding concurrent renal disease, Dr. Katarina Hazuchova and PD Dr. Florian Zeugswetter are in agreement: if euthyroidism has been achieved

using antithyroid medication and there is no severe azotaemia at that point (creatinine within IRIS stage 2), the cat can be a good candidate for radioiodine therapy. It is very rare for azotaemia to worsen significantly after RIT. However, it is essential to ensure that hypothyroidism does not develop. If azotaemia or clinical signs of hypothyroidism occur alongside low T4 concentrations, levothyroxine supplementation should be initiated.

Monitoring: when should follow-up take place after starting therapy – and what should the final T4 target be? Dr. Katarina Hazuchova recommends the first recheck after 3–4 weeks in stable patients: full blood count, renal parameters, T4, and clinical assessment of symptoms. After another 3–4 weeks, another check is performed; once stabilised, monitoring every 3–4 months is usually sufficient.

Prof. Dr. Felicitas Boretti takes a different approach for cats that are already azotaemic at diagnosis, recommending earlier follow-up after 1–2 weeks.

She also advises measuring TSH to avoid missing iatrogenic hypothyroidism (it should not rise above the reference interval). PD Dr. Florian Zeugswetter defines a different target range for T4 in azotaemic cats: in non-azotaemic cats, the goal is a T4 concentration in the middle of the reference range or slightly below. In azotaemic patients, however, T4 may remain in the upper reference range.

Prof. Dr. Boretti confirms that the timing of blood sampling in relation to tablet administration does not matter. Fluctuations in T4 concentrations during antithyroid therapy are well known, but they do not follow a circadian rhythm and are not related to the timing of medication. However, she emphasises that T4 should be measured in an external, quality-assured laboratory. In-house devices are less reliable and may deviate even within clinically relevant concentration ranges, which can lead to misinterpretation.

The other experts strongly agree with this assessment.

Finally, the topic of side effects is briefly addressed. Dr. Katarina Hazuchova distinguishes between side effects that can often be “waited out” and those that require immediate action. The most common are gastrointestinal side effects – unpleasant for owners, but usually temporary. They should therefore be discussed proactively from the start.

Mild increases in liver enzymes may also occur; these are rare and can normalise despite ongoing therapy. In contrast, severe pruritus and, in particular, blood dyscrasias such as haemolytic anaemia, neutropenia/leukopenia, or thrombocytopenia are critical – in these cases, antithyroid medication must be discontinued.

Such severe adverse effects, PD Dr. Astrid Wehner notes, most often occur within the first month and frequently within the first three months. The choice of oral formulation does not influence their frequency. Dermal preparations, however, may be associated with fewer gastrointestinal side effects.

Prof. Dr. Felicitas Boretti concludes on a reassuring note: even blood count abnormalities are usually reversible after discontinuation of antithyroid drugs. Modern, well-adjustable veterinary preparations allow for optimal control of T4 concentrations, meaning that severe side effects are now seen significantly less often.

Jennifer von Luckner, Ruth Klein

Il cane – La proteina C-reattiva (CRP) come standard di riferimento

Essendo una proteina di fase acuta (APP) fondamentale, la CRP aumenta di 50-100 volte entro 4-24 ore dopo l’esposizione a uno stimolo scatenante, raggiunge il suo picco massimo dopo 1-2 giorni e diminuisce rapidamente con un trattamento efficace. Grazie a questo andamento dinamico, la CRP è ideale per la diagnosi precoce, il monitoraggio della progressione della malattia e la valutazione del successo del trattamento.

Livelli elevati di CRP si riscontrano in una varietà di processi infiammatori e immunomediati, tra cui infezioni batteriche, infestazioni parassitarie,  malattie autoimmuni, neoplasie e alterazioni post-traumatiche o post-operatorie.
Nei cani con infezione acuta da Babesia canis, esiste una chiara correlazione tra CRP e gravità clinica e parametri ematologici.

Nell’ambito della gestione antimicrobica, è stato dimostrato che gli antibiotici possono essere sospesi una volta ottenuto un miglioramento clinico e la normalizzazione della concentrazione di CRP, riducendo significativamente la durata della terapia per molte patologie.

Nelle micosi sistemiche, come la coccidioidomicosi polmonare, la CRP in combinazione con l’Hp ha mostrato un valore predittivo per la remissione.

Tuttavia, la CRP può risultare elevata anche in assenza di una causa infiammatoria, ad esempio durante un’intensa attività fisica o in gravidanza; per questo motivo, l’interpretazione deve essere contestualizzata.

 

Tabella 1: Panoramica delle variazioni delle proteine di fase acuta e della conta leucocitaria in funzione del tempo trascorso dallo stimolo infiammatorio in cani e gatti.

Tempo da stimolo infiammatorio	Cane – ad es. CRP, SAA	Cane – conta leucocitaria	Gatto – ad es. SAA, AGP	Gatto – conta leucocitaria
0–6 h	inizialmente lieve aumento (la sintesi nel fegato inizia dopo alcune ore).	perlopiù ancora entro l’intervallo di riferimento	inizialmente lieve aumento	perlopiù ancora entro l’intervallo di riferimento
6–12 h	aumento significativo misurabile	tendenza iniziale verso un possibile aumento, spesso ancora al limite	aumento significativo misurabile	tendenza iniziale verso un possibile aumento, spesso ancora al limite
12–24 h	forte aumento, valori per lo più chiaramente patologici	leucocitosi/-penia è ora frequentemente visibile	forte aumento, chiaramente patologico	leucocitosi frequente, a volte leucogramma da stress
24–48 h	Picco APR – massima concentrazione	ulteriore aumento/plateau dei leucociti	Picco APR	ulteriore aumento/plateau dei leucociti
2–5 gg	declino incipiente, quando infiammazione sotto controllo	leucociti spesso ancora elevati, diminuiscono lentamente	declino con miglioramento clinico	leucociti spesso sono ancora alterati, normalizzano lentamente
> 5 gg	ritorno a valori di riferimento o a valori prossimi a quelli di riferimento in guarigione da infiammazione	normalizzazione, ma può richiedere più tempo a causa di processi cronici	simile al cane	simile al cane

APR = reazione di fase acuta, CRP = proteina C-reattiva, SAA = amiloide sierica A, AGP = alfa-1-glicoproteina acida

 

Indice di Fase Acuta (API) – marcatore combinato

La ricerca attuale combina i marcatori APP positivi (CRP, Hp) e negativi (albumina, eventualmente PON-1) in un Indice di Fase Acuta (API). Questo indice riflette l’attività infiammatoria complessiva.
I cani con tumori maligni e valori elevati di API presentavano una prognosi significativamente peggiore.

Nelle malattie infiammatorie croniche, ad esempio la leishmaniosi canina, i livelli di CRP e Hp rimangono costantemente elevati, mentre quelli di albumina e transferrina spesso diminuiscono. Le variazioni dell’API sono strettamente correlate alla risposta al trattamento e all’attività della malattia. Valori persistentemente elevati indicano attività residua, coinfezioni o fallimento del trattamento.

 

 

Il gatto – SAA e AGP sotto i riflettori

Nei gatti, la dinamica e il significato delle APP differiscono considerevolmente da quelli dei cani. La SAA è la principale APP più importante, mentre l’AGP è particolarmente importante per la diagnosi di FIP.

 

Siero amiloide A (SAA)

La SAA reagisce molto precocemente e con elevata sensibilità, raggiungendo rapidamente concentrazioni elevate ed è quindi adatta sia per la diagnosi precoce che per la valutazione prognostica. Un rapido declino indica una buona risposta alla terapia, mentre un valore stagnante suggerisce un’infiammazione persistente o un’infezione secondaria. I moderni test turbidimetrici con anticorpi monoclonali offrono un’elevata precisione diagnostica. Ulteriori studi dimostrano l’utilità della SAA, in particolare nelle infezioni batteriche come la pielonefrite.

 

Glicoproteina alfa-1-acida (AGP)

L’AGP è una proteina di fase acuta (APP) che aumenta moderatamente e ha un’elevata rilevanza clinica per la FIP (Fig. 1). Livelli elevati di AGP nel siero supportano la diagnosi sospetta in combinazione con altri riscontri. L’AGP mostra variazioni particolarmente dinamiche durante la terapia antivirale.

 

Aptoglobina (Hp)

Nei cani e nei gatti, l’Hp è una delle proteine di fase acuta a moderata espressione sintetizzate nel fegato. La sua funzione biologica principale è il legame ad alta affinità con l’emoglobina libera (Hb) proveniente dagli eritrociti lisati, riducendo così il danno ossidativo ai tessuti e prevenendo la perdita di ferro legato all’Hb. Durante i processi infiammatori acuti, entrambe le specie mostrano un aumento della concentrazione di aptoglobina significativamente meno pronunciato e più ritardato rispetto alle principali proteine di fase acuta come la SAA o la CRP. Come in altri mammiferi, l’emolisi intravascolare può portare a una diminuzione della concentrazione di aptoglobina perché la proteina viene rapidamente consumata legando grandi quantità di emoglobina libera.

 

Proteine di fase acuta negative

Albumina
L’albumina diminuisce a causa della ridistribuzione degli aminoacidi per la sintesi delle proteine di fase acuta positive e a causa dell’aumento della permeabilità capillare. È un indicatore prezioso di infiammazione sistemica, ma deve essere valutata tenendo conto di idratazione, perdita di proteine e funzionalità epatica. Nei cani, l’albumina è inclusa nel parametro API.

Transferrina
La transferrina, una proteina di trasporto che lega il ferro, diminuisce durante la fase di risposta parziale acuta (APR) per ridurre la disponibilità di ferro per i microrganismi. Nei cani, è stata osservata una marcata diminuzione della transferrina nelle infezioni batteriche. Una diminuzione significativa è stata documentata anche nei gatti con infiammazione cronica.

 

Proteine di fase acuta nella FIP

La peritonite infettiva felina (FIP) è una malattia infiammatoria solitamente associata a un aumento delle proteine di fase acuta.
Studi hanno dimostrato che la determinazione dell’AGP nel liquido essudativo è il metodo più significativo per differenziare i gatti con FIP da quelli senza. Sono stati definiti diversi intervalli di cut-off con sensibilità/specificità variabili (Tabella 2). Alcuni di questi intervalli di cut-off, ad esempio, in Helfer-Hungerbühler et al. (AGP > 2927), mostrano un’elevata specificità (97%) e possono quindi essere altamente indicativi di FIP.
Tuttavia, a causa della sua sensibilità relativamente bassa (54%), quasi la metà dei gatti affetti da FIP potrebbe non essere diagnosticata. Inoltre, va considerato che le APP possono aumentare anche in altre patologie. I gatti con addome settico e anche i gatti con neoplasie disseminate spesso mostrano concentrazioni di AGP altrettanto elevate quanto quelle dei gatti affetti da FIP. Pertanto, è importante eseguire un esame citologico e batteriologico supplementare per escludere diagnosi differenziali.

La sola misurazione dell’AGP non è quindi sufficiente per una diagnosi definitiva, ma rappresenta piuttosto uno dei tanti elementi necessari per giungere a una diagnosi.
L’AGP può anche svolgere un ruolo importante nel monitoraggio della terapia dei gatti affetti da FIP. Durante il trattamento, l’AGP diminuisce costantemente, ma più lentamente rispetto alla SAA. Ciò è presumibilmente dovuto alla maggiore emivita dell’AGP, motivo per cui la concentrazione di AGP al secondo giorno di terapia per la FIP può essere superiore rispetto a prima dell’inizio del trattamento. Una diminuzione significativa dell’AGP è stata osservata a partire dal settimo giorno dopo l’inizio della terapia. Entro il 28° giorno al più tardi, l’AGP era tornata entro i valori normali in quasi tutti i gatti (Helfer-Hungerbuehler 2024: (10) – 17/18 gatti e Zuzzi-Krebitz 2024: 37/39 gatti) e può quindi essere utilizzata come un buon parametro per monitorare il successo del trattamento.
Per quanto riguarda la SAA, una diminuzione significativa era già evidente al giorno 2 e la maggior parte dei gatti ha mostrato nuovamente concentrazioni di SAA (quasi) normali entro 4-7 giorni.
Addie et al. (2022) utilizzano l’AGP come marcatore per differenziare tra remissione e guarigione. La guarigione si riferisce alla completa cura della FIP, mentre la remissione è definita come uno stadio intermedio tra la guarigione e la morte, che comporta ancora il rischio di recidiva. I gatti completamente guariti mostravano livelli di AGP entro i valori normali, mentre i gatti in remissione mostravano livelli elevati di AGP. L’aumento dell’AGP potrebbe quindi indicare anche una potenziale recidiva di FIP.

 

Tabella 2: Panoramica delle pubblicazioni attuali sull’uso dell’AGP. Valori medi misurati (incluso l’intervallo) nei gatti con FIP rispetto ai gatti senza FIP, valori di cut-off definiti e relativa sensibilità e specificità.

Studio	Proteina di fase acuta	Valore medio in gatti con FIP (range)	Valore medio in gatti senza FIP (range)		cut-Off	Sensibilità (%)	Specificità (%)
	Siero
Hazuchova 2017	AGP (µg/ml)	2900 (960-5040)	690 (120-4500)		2260	85	90
	SAA (µg/ml)	98,5 (1,3-163,4)	7,6 (0,1-163,8)		97,3	55	87
	Hp (mg/ml)	2,0 (2,0-9,0)	1,8 (0,0-2,0)		2,0	55	82
	Versamento
	AGP (µg/ml)	2570 (1300-5760)	480 (190-3800)		1550	93	93
	SAA (µg/ml)	80,4 (0,1-207,4)	0,1 (0,1-182,7)		43,6	71	91
	Hp (mg/ml)	2,2 (0,1-9,3)	0,8 (0,1-2,5)		2,1	79	87
	Siero
Helfer- Hungerbuehler 2024	AGP (µg/ml)	2954 (200-5861)	sano	malato	2531	61	79
			235 (78-616)	1734 (305-3449)	2927	54	97
	Versamento
	AGP (µg/ml)	2425 (343-5611)	560 (83-3950)		1686	71	89
	Siero
Romanelli 2024	AGP (µg/ml)	1986 (405-4428)	296 (165-4254)		707	80	80
					>4099	–	100
					<438	100	–
	Versamento
	AGP (µg/ml)	1717 (549-3166)	233 (103-4099)		990	75	73
					>4254	–	100
					<296	100	–

 

 

Dott.ssa. Ruth Klein, Katharina Buchta