ELISA Test

Indications, advantages, reference concentration, specificity and sensitivity, method of detection, sample material, short protocoll

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Indications

Diagnosis or exclusion of exocrine pancreatic insufficiency caused by, for example, pancreatic acinar atrophy, pancreatic tumor, chronic pancreatitis, parasites or stones.

Advantages

In contrast to conventional laboratory parameters of canine pancreatic exocrine function, the determination of pancreatic elastase 1 has the following advantages:

  • The test is non-radioactive. An isotopic laboratory is not required.
  • In contrast to the cTLI test, canine elastase 1 measurement has the advantage that acute inflammatory episodes do not result in false normal values.
  • E1 is absolutely pancreas-specific.
  • Since E1 is stable during intestinal transit, the faecal elastase 1 concentration reflects the secretory capacity of the pancreas (diagnosis or exclusion of pancreatic exocrine insufficiency).
  • Digestive enzyme substitution therapy has no influence on the determination of E1. The monoclonal antibodies used in the test are monospecific for canine elastase 1, therefore recognizing only elastase 1 of canine origin. There is no cross-reaction with bovine or porcine elastases, contained in the preparations used for enzyme substitution therapy.
  • Substitution therapy does not need to be discontinued 5 days before measurement, in contrast to the determination of faecal chymotrypsin activity.
  • There is no need for a 12-hour starvation period, in contrast to the necessary preparation before the cTLI test.
  • The Quick-Prep™ stool sample extraction system is available also (Cat.-No. 09-Quick).

Reference concentration

Values above 40 µg/g faeces indicate normal pancreatic exocrine function.
Values below 10 µg/g faeces indicate a severe pancreatic exocrine insufficiency.
Borderline values (“grey zone”) of 10 – 40 µg/g faeces suggest the beginning of pancreatic exocrine insufficiency.

High specificity and sensitivity

Determined for clinically manifest pancreatic insufficiency (Cut off < 10 µg/g)

Specificity: 92% Sensitivity: 95%

Method of detection

Sandwich ELISA (96-well format) based on two monoclonal antibodies highly specific for
canine pancreatic elastase 1.

Sample material

  • A single pea-sized stool sample is sufficient.
  • High stability of pancreatic elastase 1 allows time for convenient mailing.
  • No need to interrupt digestive enzyme substitution therapy.

Short protocol for the experienced user
Important: The short protocol is not a substitute for the detailed protocol given in the instruction manual!

  • Prepare the sample-/washing buffer and the extraction buffer
  • Extract and homogenize faeces
  • Dilute faeces extract in sample-/washing buffer
  • Pipette 50 µl blank, standards, control and samples in duplicate into the ELISA-strips
  • Incubate 60 minutes at room temperature
  • Wash
  • Add 50 µl anti E1-bio (1:100)
  • Incubate 30 minutes at room temperature
  • Wash
  • Add 50 µl POD-Streptavidin-Complex (ready-to-use)
  • Incubate 30 minutes at room temperature (in the dark)
  • Wash
  • Add 100 µl TMB substrate solution (ready-to-use)
  • Incubate 20 minutes at room temperature (in the dark)
  • Add 100 µl stop solution (ready-to-use)
  • Read plate at OD 450 or OD 450 – OD 620 between 5 and 30 minutes after addition of the stop solution
  • Evaluate with standard curve using a log-log scale