Determining the IgG concentrations in bovine colostrum and calf sera with a novel enzymatic assay

Abstract

BackgroundImmune protection in newborn calves relies on a combination of the timing, volume and quality of colostrum consumed by the calf after birth. Poor quality colostrum with inadequate immunoglobulin concentration contributes to failed transfer of passive immunity in calves, leading to higher calf morbidity and mortality. Therefore, estimating colostrum quality and ensuring the transfer of passive immunity on farm is of critical importance. Currently, there are no on-farm tools that directly measure immunoglobulin content in colostrum or serum. The aim of this study was to apply a novel molecular assay, split trehalase immunoglobulin G assay (STIGA), to directly estimate immunoglobulin content in dairy and beef colostrum and calf sera, and to examine its potential to be developed as on-farm test. The STIGA is based on a split version of trehalase TreA, an enzyme that converts trehalose into glucose, enabling the use of a common glucometer for signal detection. In a first study, 60 dairy and 64 beef colostrum and 83 dairy and 84 beef calf sera samples were tested with STIGA, and the resulting glucose production was measured and compared with radial immunodiffusion, the standard method for measuring immunoglobulin concentrations.ResultsPearson correlation coefficients between the methods were determined and the sensitivity, specificity, and accuracy of the test were calculated for different colostrum quality and failed transfer of passive immunity cut-off points. The correlations of the STIGA measured by colorimetric enzymatic reaction compared to radial immunodiffusion for dairy and beef colostrum were 0.72 and 0.73, respectively, whereas the correlations for dairy and beef sera were 0.9 and 0.85, respectively. Next, STIGA was tested in a blinded study with fresh colostrum and serum samples where the correlation coefficient was 0.93 and 0.94, respectively. Furthermore, the performance of STIGA followed by glucometer readings resulted in correlations with radial immunodiffusion of 0.7 and 0.85 for dairy and beef colostrum and 0.94 and 0.83 for dairy and beef calf serum.ConclusionsA split TreA assay was validated for measurement of the immunoglobulin content of colostrum and calf sera using both a lab-based format and in a more user-friendly format compatible with on-farm testing.

Detecting total immunoglobulins in diverse animal species with a novel split enzymatic assay

“…The reaction was incubated for 90 min at 25 °C. Absorbance (OD) was measured after 90 min in EnSpire multimode plate reader at 450 nm (EnSpire, PerkinElmer, Waltham, MA) [24, 25].…”

“…Recently, a novel detection assay (named Split TreA Immunoglobulin G Assay or STIGA) was developed to detect antibodies in bovine colostrum and calf serum [24]. The assay is based on split trehalase technology [25].…”

“…Specifically, TreA dimerization efficacy decreases with the concentration of the analyte (IgG) [25]. In the previous study [24], split TreA carrying protein G (STIGA) was applied to quantify the amount of bovine IgGs in colostrum and calf serum. There, the optimal concentration of reagents for this type of assay was established and the new test format (92-well plate format) was developed.…”

Alimentación: factor estratégico durante la crianza artificial de terneros provenientes de lecherías

“…Debido a que la IgG es la que determina la calidad del calostro, es necesario realizar su medición. Para ello, se han utilizado técnicas como, el uso de calostrómetro (Bartier et al, 2015;Angulo et al, 2015) y la refractometría (Bielmann et al, 2010), que son consideradas de tipo indirecto y pueden realizarse en campo; mientras que, existen otras técnicas de tipo directo, que se realizan en laboratorio, como la inmunodifusión radial (RID) (Gelsinger et al, 2015), ELISA (Dunn et al, 2017), la espectroscopía de transmisión infrarroja (IRS) (Elsohaby et al, 2018) y recientemente, nuevos ensayos enzimáticos (Drikic et al, 2018) (Cuadro 2). Siendo estas últimas, las técnicas menos comunes y más recientemente exploradas.…”

“…Referente a ensayos enzimáticos, se probó una técnica molecular basada en la medición directa de IgG tanto en suero sanguíneo como en calostro, con una prueba denominada ensayo de inmunoglobulina G de trehalasa dividida (STIGA). Los autores hallaron una alta sensibilidad, especificidad y exactitud de la técnica comparada con otros métodos como RID, validando dicho método como otra alternativa para medir IgG (Drikic et al, 2018).…”