Feline pancreatic lipase: purification and validation of a clinically significant radioimmunoassay for the diagnosis of feline pancreatitis
Pancreatic lipase was purified from feline pancreatic tissue by delipidation, anion-exchange chromatography, size-exclusion chromatography, and cation-exchange chromatography. Antiserum against fPL was raised in rabbits. Tracer was produced by iodination (125I) of fPL using the chloramine T method. An RIA was established and validated by determination of sensitivity, dilutional parallelism, spiking recovery, intraassay variability, and inter-assay variability. A control range for fPLI in cat serum was established from 30 clinically healthy cats using the central 95th percentile.
The sensitivity of the fPLI assay was 1.2 ?g/L. Observed to expected ratios for serial dilutions ranged from 58.0 to 164.3% for 4 different serum samples at dilutions of 1 in 2, 1 in 4, and 1 in 8. Observed to expected ratios for spiking recovery ranged from 76.0 to 156.5% for 4 different serum samples and 6 different spiking concentrations. Coefficients of variation for intra-assay variability for 4 different serum samples were 10.1, 4.5, 2.2, and 3.9%. Coefficients of variation for inter-assay variability for 4 different serum samples were 24.4, 15.8, 16.6, and 21.3%. The control range for serum fPLI concentration was established as 1.2 to 3.8 ?g/L.
All of the objectives outlined above were successfully met, leading to the development of an RIA for the measurement of fPLI in cat serum. The RIA for fPLI described here is sufficiently accurate and precise, but has a limited linearity and reproducibility in the lower and higher end of the working range.
Advisor:Steiner, Jörg M.; Williams, David A.; Berghman, Luc R.
School:Texas A&M University
School Location:USA - Texas
Source Type:Master's Thesis
Keywords:feline pancreatitis lipase pancreatic radioimmunoassay ria
Date of Publication:12/01/2003