MEGACOR FASTest C perfringens Toxin User Manual

MEGACOR FASTest C perfringens Toxin

Test-kit for the qualitative detection of Clostridium perfringens enterotoxin (CPE) in the feces of the dog, cat, goat and sheep lamb, calf, foal and piglet

Supplied Exclusively To The UK Veterinary Market By
Vetlab Supplies Ltd
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INFORMATION ON THE TEST-KIT

TEST-KIT COMPONENTS

1 test-kit FASTest® C. perfringens Toxin contains:

• 2 or 10 dipsticks coated with monoclonal antibodies
• 2 or 10 sample tubes with 2.0 ml buffer diluent each
• 1 instruction for use

LIABILITY
The entire risk due to the performance of this product is assumed by the purchaser. The manufacturer shall not be liable for indirect, special or consequential damages of any kind resulting from the use of this product.

INTRODUCTION

The gram-positive anaerobic bacterium Clostridium perfringens belongs to the physiological intestinal flora of many mammals and is facultative pathogenic. Inconvenient endogenous (other basic diseases, diarrhea pathogens, antibiotic therapies with massive reduction of intestinal fl ora etc.) and exogenous (farming condi-tions, extreme changes of the food, stress etc.) factors can lead to increased pathogenicity of C. perfringens. Next to its ability to form extremely infectious and stable spores, the formation of lethal toxins is crucial for its pathogenicity. The classifi cation into the various types (A–E) is only due to the toxin formation.
These toxins can cause extremely variable (mild to lethal progression forms) failures of the intestinal water and electrolyte balance in different species like goat, sheep (e. g. dysentery of lambs: type B; pulpy kidney disease: type D), cattle (hemorrhagic enteritis: type A–E), foal (hemorrhagic necrotizing enteritis: type A & C) and piglet (e. g. serous-catarrhal enteritis: type A, necrotizing enteritis: type C).
In the dog, especially serotype A occurs, producing 2 main toxins (toxin Alpha [α] and a Clostridia enterotoxin [CPE]), rarer serotype B (toxin Beta [β]). Both C. perfringens and its CPE can be detected also in healthy dog’s feces. CPE can be detected more often in dogs with diarrhea compared to healthy dogs. CPE is more frequent in dogs with diarrhea (hemorrhagic gastroenteritis, acute or chronic diarrhea, enterotoxaemia) than in healthy dogs. For cats, to date reliable literature data concerning prevalence and clinical relevance are missing.
Only by detection of C. perfringens in the feces, a disease caused by Clostridia is not diagnosable. In a study in Switzerland, 54 % of the C. perfringens isolates showed a reduced sensitivity towards metronidazole or 18 % towards tetracycline. Because there is a general risk of resistance formation, it is recommended to identify the triggering pathogen in principle. By its high sensitivity and specifi ci-ty, the use of FASTest® C. perfringens Toxin allows the veterinarian a rapid aetiological on-site diagnosis of a C. perfringens infection, the quick initiation of therapy as well as of necessary quarantine and prophylaxis measures.

INFORMATION ON THE SPECIMEN MATERIAL

Due to the normally inhomogeneous or nest-like dissemination of antigens in the feces, the specimen material has to be mixed up homogeneously (spatula, vortex-mixer) before sampling.
For the test, the required amount of feces as described in issue 4b / Specimen collection and preparation, is needed. The amount depends on the consistency of the sample. Use the attached spoon.
Non-cooled (15–25 °C), the sample should be tested within 4 hours! At 2–8 °C, the sample can be stored up to 4 days, permanently at minimum −20 °C.
Keep in mind that the sample material, as well as all used test-kit components, should have reached room temperature at the time of application.
Endogenous and exogenous interfering substances of the sample (e. g. proteases, mucosa components, blood, but also viscosity, pH-value as well as grass and cat litter) can cause interferences (matrix effects) that can infl uence the target measurement. These can lead to an impaired LF and/or unspecifi c reactions on TL and CL.

SPECIMEN COLLECTION AND PREPARATION

• Open the sample tube with the buffer diluent.
• Mix the feces sample homogeneously (applicator, vortexer). Then mix the required sample volume (compact: 1 level spoon, pulpy: 2 level spoons, fluid-watery: 3 lev-el spoons of feces) steadily into the buffer diluent (fi g.1).

• Close sample tube tightly and rotate it easily to get the mixture as homogeneous as possible (fi g.2).
• For sedimentation of gross feces particles place the sample tube on a fl at and horizontal surface for 1–5 minutes.

TEST PROCEDURE

1. Remove the dipstick from its foil pouch shortly before use.
2. Introduce the dipstick vertically and with the arrows pointing downwards into the sample tube for at least 1 minute. The liquid level (meniscus!) must not exceed the blue horizontal line below the blue arrowheads (fi g.3).
3. Remove the dipstick from sample tube soonest the sam-ple-buffer mixture (SBM) has reached the CL. If so, the pink-purple CL will appear slowly but surely (fi g.4 / 5). If the CL will not appear after 5–10 minutes, a new SBM must be prepared and sedimented for at least 5 minutes. The dipstick must be held only in the supernatant until the LF has reached the CL (see also 7. Precautions for users*).
4. Place the dipstick on a fl at and horizontal surface for incubation.

READING OF THE TEST RESULT

Read the test result 5 (max. 10) minutes. Positive test results may be observed earlier, depending on the concentration of CPE in the sample.

POSITIVE TEST RESULT (fi g.4)
A pink-purple TEST line of any intensity (varying from very weak to strongly intensive) and a pink-purple CONTROL line appear.

NEGATIVE TEST RESULT (fi g.5)
Only a pink-purple CONTROL line appears. This line indicates, irrespective of its intensity, that the test has been performed properly.

INVALID TEST RESULT
No CONTROL line visible. The test should be repeated using a new dipstick *.

PRECAUTIONS FOR USERS

• The guidelines for working in medical laboratories must be observed. It is recommended to wear disposable gloves and other personal protective equipment (protec-tive clothing, possibly a face mask). Wash and disinfect hands after completing the test.
• Label sample material and associated sample tube to ensure a precise assignment.
• Use a new sample tube and a new dipstick for each sample.
•  The buffer diluent contains low concentrations of toxic sodium azide as a preservative, therefore avoid skin/eye contact and/or ingestion.
• The sample material must be seen as potentially infectious and disposed of accordingly, together with the used test-kit components.
• To avoid an application error / external infl uence (e. g. too much sample material, too short sedimentation time, com-ponents in the faeces that clog the pores of the suction pad), the test can be repeated. Use a new dipstick and carefully observe the sample preparation. It is advisable to only hold the dipstick in the supernatant when repeating the test until the LF has reached the CL.

TEST PRINCIPLE

The FASTest® C. perfringens Toxin is based on latest rapid immunochromatographic technique.
The Clostridium perfringens enterotoxin (CPE) in the feces sample will react in the conjugate pad area with mobile monoclonal anti-CPE antibodies (anti-CPE mAbs), which are bound to gold particles. Migrating (“lateral fl ow”, LF) along the nitrocellulose membrane, this specifi c antigen-antibody complexes are bound by fixed anti-CPE mAbs producing a pink-purple TEST line (TL). These anti-CPE mAbs guarantee a high level of specifi city for the aetiological detection of Clostridium perfringens enterotoxin. The intensity or width of TL depends on the concentration of Clostridium perfringens enterotoxin in the tested sample.
A correct test procedure will be indicated by a second, pink-purple CONTROL line (CL).

INFORMATION FOR THE INTERPRETATION

• The interpretation of the test result should always be based on anamnestic and clinical data as well as the therapy and prophylaxis possibilities.
• Any non-described color or contour variation of TL and CL within the indicated incubation time or after more than 10 minutes (e. g. greyish, shadow-like lines) has to be considered as an unspecific reaction and therefore as a negative test result.
• TL can vary both in intensity (from weak to intense pink-purple) and width. Therefore, any pink-purple line appearing within the required incubation time is to be
  interpreted as a positive test result.

References

• MEGACOR Diagnostik GmbH – Veterinary in vitro diagnostics
• Wholesale Vet Supplies & Products for Practices & Labs | Vet Supplies Online UK

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