Emerging technologies for the assessment of bovine immunoglobulins in biofluids

Abstract

Measurement of bovine serum and colostrum immunoglobulin G (IgG) concentration is critical for colostrum and calf health monitoring in order to determine the colostrum quality and failure of transfer of passive immunity (FTPI), which is considered the main reason for increased morbidity and mortality rates in newborn calves. Several qualitative and quantitative assays are available, but radial immunodiffusion assay is acknowledged as the reference method. However, it is expensive and takes a long time to obtain the results, which prevents the identification of calves with FTPI prior to gut closure. As a consequence, there is a demand for rapid, accurate and inexpensive diagnostic assays for IgG. The objectives of this thesis were: (1) to develop and validate an infrared (IR) spectroscopy based assay for quantification of bovine serum and colostrum IgG concentration, and (2) to validate a novel and rapid on-farm tool for detection of FTPI in dairy calves and for assessing colostrum quality of dairy cows. Infrared spectroscopy has recently emerged as a powerful, reagent-free diagnostic tool for the quantitative characterization of biological fluids in human and veterinary medicine. We set out to develop a quantitative assay based on IR spectroscopy to measure bovine serum and colostrum IgG concentration and to compare the values with that of the RID assay. Although there are a number of IR spectroscopic sampling techniques, in this research, transmission-IR and attenuated total reflectance (ATR-IR) spectroscopy were used. For quantification of bovine serum IgG concentration, transmission-IR and ATR-IR assays were developed using 250 serum samples collected from calves. The IgG concentration measured by both assays showed excellent correlation with RID-measured IgG. Also, both IR-based assays showed potential for detection of FTPI with good to excellent sensitivity, specificity and accuracy. The transmission-IR assay showed slightly higher precision than the ATR-IR assay. However, the ATR-IR assay is more appropriate for farm and veterinary clinic use. For quantification of colostral IgG concentration, a transmission-IR assay was developed using 251 colostrum samples. The IgG measured by the IR assay showed excellent levels of agreement with the RID assay. The results suggest that IR spectroscopy may be a useful method for colostrum monitoring programs. Evaluation of an initial version of ZAPvet Bovine IgG test for detection of calves with and without FTPI revealed that the ZAPvet test is relatively sensitive and would be acceptable as an initial screening test for diagnosis of FTPI in dairy calves. However, the low specificity of ZAPvet test would result in over prediction of FTPI incidence, which could result in unnecessary interventions for calves with adequate transfer of passive immunity. Validation of refractometers, either digital Brix (Atago Co. Ltd; WA) or optical STP (Westover RHC-200ATC handheld refractometer, Woodinville, WA), for detection of FTPI in 202 dairy calves, and digital Brix (Atago Co. Ltd; WA) or optical Brix (model 300001; SPER Scientific, Scottsdale, AZ), for assessing of quality of 251 colostrum samples, revealed that the digital and optical refractometers have good potential for being a useful and practical on-farm management tools to be included in colostrum and calf health monitoring program on dairy operations. Furthermore, the two refractometers performed similarly for detection of FTPI in dairy calves and for assessing of colostrum quality of dairy cows with cut-points slightly higher than that reported in recent studies